From KMB1904 <@t> aol.com Sun Oct 2 07:07:28 2005 From: KMB1904 <@t> aol.com (KMB1904@aol.com) Date: Sun Oct 2 07:07:48 2005 Subject: [Histonet] immuno exam. Message-ID: Does anyone know where I can get learning materials for the immuno exam for the ASCP?? Thanks From c.m.vanderloos <@t> amc.uva.nl Mon Oct 3 01:43:19 2005 From: c.m.vanderloos <@t> amc.uva.nl (C.M. van der Loos) Date: Mon Oct 3 01:43:45 2005 Subject: [Histonet] RE: Tissue fixation help Message-ID: Angela, Long time ago we thawed a frozen tissue sample directly in (room temp.) NBF (24-48 hs). Morphology was kept surprisingly well. Sorry, at that time phosphorylated antibodies were not even existant.... Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands phone: +31 20 5665631 Date: Fri, 30 Sep 2005 08:25:24 -0400 From: Angela McNabola Subject: [Histonet] Tissue fixation help To: [1]Histonet@lists.utsouthwestern.edu Good Morning, Does anybody on the histonet have any experience with taking tissue that has been snap frozen in minus 80, and then "thawed" out and formalin fixed and paraffin embedded? I won't get into the reasons why we have to do this, but does anyone have a protocol (or just slowly thaw it out and fix it??). How did your IHC turn out, especially when using phosphorlyated antibodies? Did you get good morphology of your tissue? thanks in advance! Angela Angela McNabola, MS HT(ASCP)SLS, QIHC Bayer Healthcare 400 Morgan Lane West Haven, CT 06516 203-812-5001 angela.mcnabola.b@bayer.com References 1. mailto:Histonet@lists.utsouthwestern.edu From brucea <@t> unimelb.edu.au Mon Oct 3 02:26:37 2005 From: brucea <@t> unimelb.edu.au (Bruce Abaloz) Date: Mon Oct 3 02:27:09 2005 Subject: [Histonet] RE: Tissue fixation help In-Reply-To: References: Message-ID: > Hi Angela, > >we periodically thaw tissue from the -80C freezer in room temp. 10% >NBF & get very good >morphology of our tissue - same as Chris van der Loos said he does. >IHC works fine but no experience with phosphorlyated antibodies? > > > -- BRUCE ABALOZ PH:61383446282 HISTOLOGIST FAX:61383447909 DEPT.of ZOOLOGY EMAIL: brucea@unimelb.edu.au THE UNIVERSITY Of MELBOURNE. VICTORIA.AUSTRALIA 3010 http://www.zoology.unimelb.edu.au/ Nobody Can Make You Feel Inferior Without YOUR Permission - Eleanor Roosevelt DANCE LIKE NO-ONE'S WATCHING Q: What's a specimen? 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From carl.hobbs <@t> kcl.ac.uk Mon Oct 3 02:26:13 2005 From: carl.hobbs <@t> kcl.ac.uk (Carl Hobbs) Date: Mon Oct 3 02:27:12 2005 Subject: [Histonet] re JKiernan APES treatment of slides Message-ID: <000e01c5c7eb$bc5890d0$112b5c9f@Carlos> Thank you very much for your reiteration of the procedure for charging > slides, using APES, Mr Kiernan. I would be grateful for your comments > regarding: > 1. Do you know if the "Superfrost plus" slides that I buy from VWR are > prepared using the same/similar process? > 2. I use a similar method to your posted one ; it differs only in that I > rinse the > slides in water after the APES/acetone treatment, then rinse in acetone > to aid a fast dry; do you think that this has any deleterious effect on > the charging of the slides? > Carl From joannew <@t> bluebottle.com Mon Oct 3 04:03:50 2005 From: joannew <@t> bluebottle.com (Joanne Whitehead) Date: Mon Oct 3 04:04:14 2005 Subject: [Histonet] "empty" cryosections! Message-ID: <1128330230.4340f3f670528@bluebottle.com> First of all, many thanks to Gayle and everyone else who has offered suggestions on freezing and sectioning. I am using a Jung CM3000 cryostat with disposable blades. The machine is showing it's age - it has needed several visits from the Leica technicians lately. But I have just installed a new antiroll plate, which is a bit of help. I have tried the brush technique also, but I can see as the tissue comes across the knife it is already pulling away from the OTC, so this is not an issue of the antiroll plate. I hadn't considered variations in temperature within the machine - I will check for that next time. I wondered if it would help if the knife was colder than the tissue - anyone tried this? A couple of pellets of dry ice on the knife holder perhaps? I will definitely try embedding the tissue in OTC before freezing next time, as well as infusing with OTC before freezing. But I wonder how to make a suitable mold? Someone has suggested aluminum foil. I ususally use re-useable plastic rings if the tissue is small enough, or else cut a piece of the bulb from a disposable plastic pasteur pipette for larger pieces, which I can just cut away after freezing. Also I understand from the comments here that mixing dry ice directly into the isopentane, rather than having the isopentance suspended above liquid nitrogen will give a more consistent temperature for snap freezing - another modification I will try next time. I had already settled on -26C for sectioning, as suggested by Gayle. I need to open up the intestine for my manipulations, so infusing the tube directly with OTC is not an option for me. But I need sections only 1 cm long, so it rolls up into a reasonably compact form. I have been rolling with villi out so they will be spread apart better, but I have also seen images rolled villi in - any preferences out there? So thanks everyone, hopefully my next attempts at freezing & sectioning will go much smoother. That brings me to my next problem, which I will describe in a separate post :) From joannew <@t> bluebottle.com Mon Oct 3 04:21:50 2005 From: joannew <@t> bluebottle.com (Joanne Whitehead) Date: Mon Oct 3 04:22:00 2005 Subject: [Histonet] inconsistent DAPI staining Message-ID: <1128331310.4340f82e6ef55@bluebottle.com> This board is a great resource - I hope this problem has also been encountered by some knowledgeable people here! Sometimes I see huge variation in the DAPI signal when I view my tissue sections under the confocal microscope, and I'm wondering if this can be an artifact of poor freezing, as the sections that look the worst tend to be the ones I had trouble sectioning and show the worst morphology. At first I thought it was a problem with my mounting medium: I ordered a hard-set vectashield with DAPI, and while I was waiting for the order I borrowed a bottle of Prolong Gold from Molecular Probes. The Prolong Gold worked fine, I just had to seal the coverslips with nail polish to prevent damage to the sections. When the vectashield hard-set arrived my first sections showed really inconsistent DAPI staining - some regions nice & bright, others really weak. Generally it the the outside edge of the section that is well stained, and the interior faint. I thought perhaps the polymerizing of the mounting medium happened too quickly, so that it would harden around the tissue, which was still a bit wet from the last wash, and not distribute evenly. So next time I air dried the slides really well before mounting, but then had problems with air bubbles getting trapped in the mounting medium, and still not good staining into the middle of the sections. I talked to the Vectashield rep, who didn't have any good suggestions, but did replace the bottle with a normal non-polymerizing DAPI medium. But the problem of seeing DAPI staining only at the edges of the sections is getting worse, and have since had the same problem with the Prolong Gold too! The DAPI I use only as a counterstain to make the pictures look nice, but I wonder if the DAPI isn't evenly distributed, then perhaps the flourescence protectant is also not evenly distributed, which would undermine my quantitative analysis of the Cy3 signal I'm looking at. Yesterday's slides were the worst of all - not only was there almost no DAPI signal (expect a very tiny margin around each section), but the Cy3 signal was almost invisible too, hence the concern about the fluoroprotectant. I usually wait 2 days before analysing my slides, as the antibody seems to continue to redistribute itself, giving lower backgound over a day or 2, so I need to know that the fluorescent signal is protected. I used to put the slides immediately at -20C after mounting, but with the hard set I had to leave them at 4C so it could polymerize properly. Help! Has anyone else dealt with this problem? What is the general wisdom regarding drying (or not) the slides before mounting? Does it have any effect? The consistency of my staining has been steadily decreasing with the morphology of my sections - I hope I can solve both problems soon! Thanks very much. Joanne Institut Curie, Paris From nperson211 <@t> comcast.net Mon Oct 3 09:46:49 2005 From: nperson211 <@t> comcast.net (nperson211@comcast.net) Date: Mon Oct 3 09:47:25 2005 Subject: [Histonet] CD11b Message-ID: <100320051446.5013.43414458000DAEA6000013952200763704CECECD02019C9D0A9F02@comcast.net> Hello Histonet, Does anyone have any experience working with CD11b in FFPE rats? I would really appreciate any suggestions or comments. Thanks in advance, Nancy Lemke Research Coordinator Hermelin Brain Tumor Center Henry Ford Hospital Department of Neurosurgery Detroit, MI From gcallis <@t> montana.edu Mon Oct 3 10:03:27 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Mon Oct 3 10:04:14 2005 Subject: [Histonet] inconsistent DAPI staining In-Reply-To: <1128331310.4340f82e6ef55@bluebottle.com> References: <1128331310.4340f82e6ef55@bluebottle.com> Message-ID: <6.0.0.22.1.20051003082814.01b64690@gemini.msu.montana.edu> A couple of things here: When using Prolong Gold with DAPI, you must blot away most of the PBS before you add the drop (generally right on top of the section) and then the coverslip. Failure to stain may be a dilution factor with too much PBS. Do NOT seal with fingernail polish especially if you ever work with GFP. These were instructions directly from Tech services at Molecular Probes. To seal a coverslip for immunofluorescent staining, we discard fingernail polish out of the tidy little bottle, rinse bottle and brush with acetone, than let it completely dry and then add permanent mounting media diluted with either toluene or xylene. We love the little brush but hate the fingernail polish. We seal coverslips with diluted permanent mounting media that is NOT miscible with water. The reason is that fingernail polish contains isopropyl alcohol that leaches into the aqueous PBS under coverslip. Be careful how you store Prolong Gold with DAPI ready to use, it must be stored with dessicant AND frozen, then brought to room temperature just before use. We used an applicator stick to put a drop on from bottle to section, and try NOT to introduce any moisture back the the Prolong Gold stock. We refreeze Prolong Gold several times until it is gone, and it works perfectly. DAPI will stain DNA in either fixed or unfixed frozen sections very nicely, but I am not sure how well it stains if the tissue has been fixed a long time in formalin or paraformaldehyde - we avoid that. You did not say whether your tissues were fixed or fresh? Your failure to stain with DAPI may not be a problem with mounting media, but with some other thing you are doing to the tissue, frozen section or during staining that affects the DAPI. I would not air dry frozen sections after immunofluorescent staining, with practice and even a tiny drop of Hard Set Vectashield or Prolong Gold, you can get excellent coverage without bubbles. You can always check for freezing artifact caused by too slow a freezing method, A simple H&E stain on a frozen section from your block will tell you if you have poor snap freezing technic and getting freezing artiface. As for sectioning success, that depends on what tissue, the temperature at which you section it, thickness, sharpness of blade or knife, fixed tissue with cryoprotection. You did not elaborate on any of this. Loss of Cy3 signal can also be caused by photobleaching during overexposure to excitation light during CLSM, and if you are taking a long time to focus, etc, look at the samples, that will occur even with Antifade reagents. In general, it is a good idea to look at your sample on day of staining or very soon thereafter. I don't understand what you mean by antibody redistribution? I have not heard of this nor done it - would anyone care to elaborate. We are more worried about loss of fluorescent signal by photobleaching but in general, these special mounting medias HELP prevent this but the fluorophores can still lose fluorescence over time and light exposure. We never go to 4C to allow Prolong Gold with DAPI to set up hard nor VectaShield Hard Set either. All coverslipped sections are inside a dark folder, sitting at room temperature. Our tissue sections are are totally stained with DAPI without any of the problems you describe. At 03:21 AM 10/3/2005, you wrote: >This board is a great resource - I hope this problem has also been >encountered by some knowledgeable people here! > >Sometimes I see huge variation in the DAPI signal when I view my >tissue sections under the confocal microscope, and I'm wondering if >this can be an artifact of poor freezing, as the sections that look >the worst tend to be the ones I had trouble sectioning and show the >worst morphology. > >At first I thought it was a problem with my mounting medium: I ordered >a hard-set vectashield with DAPI, and while I was waiting for the >order I borrowed a bottle of Prolong Gold from Molecular Probes. The >Prolong Gold worked fine, I just had to seal the coverslips with nail >polish to prevent damage to the sections. When the vectashield >hard-set arrived my first sections showed really inconsistent DAPI >staining - some regions nice & bright, others really weak. Generally >it the the outside edge of the section that is well stained, and the >interior faint. I thought perhaps the polymerizing of the mounting >medium happened too quickly, so that it would harden around the >tissue, which was still a bit wet from the last wash, and not >distribute evenly. So next time I air dried the slides really well >before mounting, but then had problems with air bubbles getting >trapped in the mounting medium, and still not good staining into the >middle of the sections. > >I talked to the Vectashield rep, who didn't have any good suggestions, >but did replace the bottle with a normal non-polymerizing DAPI medium. >But the problem of seeing DAPI staining only at the edges of the >sections is getting worse, and have since had the same problem with >the Prolong Gold too! > >The DAPI I use only as a counterstain to make the pictures look nice, >but I wonder if the DAPI isn't evenly distributed, then perhaps the >flourescence protectant is also not evenly distributed, which would >undermine my quantitative analysis of the Cy3 signal I'm looking at. >Yesterday's slides were the worst of all - not only was there almost >no DAPI signal (expect a very tiny margin around each section), but >the Cy3 signal was almost invisible too, hence the concern about the >fluoroprotectant. I usually wait 2 days before analysing my slides, >as the antibody seems to continue to redistribute itself, giving >lower backgound over a day or 2, so I need to know that the >fluorescent signal is protected. I used to put the slides immediately >at -20C after mounting, but with the hard set I had to leave them at >4C so it could polymerize properly. > >Help! Has anyone else dealt with this problem? What is the general >wisdom regarding drying (or not) the slides before mounting? Does it >have any effect? The consistency of my staining has been steadily >decreasing with the morphology of my sections - I hope I can solve >both problems soon! > >Thanks very much. >Joanne >Institut Curie, Paris > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From gcallis <@t> montana.edu Mon Oct 3 10:06:10 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Mon Oct 3 10:06:28 2005 Subject: [Histonet] Re: rfp and gfp In-Reply-To: <872D009A552E7673E346EDBA@pys-aep6.pys.bris.ac.uk> References: <872D009A552E7673E346EDBA@pys-aep6.pys.bris.ac.uk> Message-ID: <6.0.0.22.1.20051003090413.01b6c448@gemini.msu.montana.edu> Patrick, Thank you for your comments and technics, now residing in my GFP file folder. Gayle Callis At 07:02 AM 10/3/2005, you wrote: >Hi Caroline and Gayle, > >Sorry for writing to you both but I think I can help a little on this >subject. I just want to echo some of Gayle's comments which are excellent >and add a few things. I currently use both eGFP and mRFP invivo. I >visualise these markers with conventional fluorescence and confocal >microscopy. I cardiac perfuse my rats with PBS (0.1M) and then perfuse >with 4% 0.1M PB formalin. I post fix for 2 hours with the same fixative >and cut 40 >mm sections using a slide microtome. I can see both eGFP and RFP equally >well. Although I will add that RFP is not as strong as eGFP and this may >be due to photobleaching/fixation. I have used both free floating and >serially mounted sections and saw no difference in >visualisation. However, if you're planning to immuno with serially >mounted sections, I would suggest permeabilising with 50% ethanol and 0.3% >triton, initially we only used 50% ethanol and saw poor >permeabilsation. You could also use anti-GFP and anti-RFP to enhance your >visualisation, I have found this has enhanced our understanding of >neuroanatomy/distribution greatly. > >I hope this has helped. > >Thanks > >Patrick > > > > > > >---------------------- >Patrick Howorth, >Dept of Physiology, >University of Bristol, >Bristol, >BS8 1TD. >+44 (0)117 33 17112 >patrick.howorth@bristol.ac.uk > > From jkiernan <@t> uwo.ca Mon Oct 3 10:06:18 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Mon Oct 3 10:06:36 2005 Subject: [Histonet] re JKiernan APES treatment of slides References: <000e01c5c7eb$bc5890d0$112b5c9f@Carlos> Message-ID: <434148EA.6E0DFA67@uwo.ca> 1. I don't know how commercial positively charged slides are made. Try asking the maker. (Good luck!) It is possible that they use an alkoxysilane other than APES to achieve a similar effect. 2. Using an acetone rinse to speed up drying should not change the APES-treated slides. The aminopropyl groups are covalently bound to the glass. They cannot be washed off, and they do not react with acetone. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ Carl Hobbs wrote: > > Thank you very much for your reiteration of the procedure for charging > > slides, using APES, Mr Kiernan. I would be grateful for your comments > > regarding: > > 1. Do you know if the "Superfrost plus" slides that I buy from VWR are > > prepared using the same/similar process? > > 2. I use a similar method to your posted one ; it differs only in that I > > rinse the > > slides in water after the APES/acetone treatment, then rinse in acetone > > to aid a fast dry; do you think that this has any deleterious effect on > > the charging of the slides? > > Carl > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From cfavara <@t> niaid.nih.gov Mon Oct 3 12:09:33 2005 From: cfavara <@t> niaid.nih.gov (Favara, Cynthia (NIH/NIAID)) Date: Mon Oct 3 12:10:18 2005 Subject: [Histonet] "empty" cryosections! Message-ID: Quick reply - I frequently cool my knife with dry ice or with that freezing spray helps sometimes. Just need to keep trying sometimes. c Cynthia Favara NIAID/NIH/RML/LPVD 903 South 4th Street Hamilton, MT 59840 406-363-9317 Disclaimer: The information in this e-mail and any of its attachments is confidential and may contain sensitive information. It should not be used by anyone who is not the original intended recipient. If you have received this e-mail in error please inform the sender and delete it from your mailbox or any other storage devices. National Institute of Allergy and Infectious Diseases shall not accept liability for any statements made that are sender's own and not expressly made on behalf of the NIAID by one of its representatives -----Original Message----- From: Joanne Whitehead [mailto:joannew@bluebottle.com] Sent: Monday, October 03, 2005 2:04 AM To: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] "empty" cryosections! First of all, many thanks to Gayle and everyone else who has offered suggestions on freezing and sectioning. I am using a Jung CM3000 cryostat with disposable blades. The machine is showing it's age - it has needed several visits from the Leica technicians lately. But I have just installed a new antiroll plate, which is a bit of help. I have tried the brush technique also, but I can see as the tissue comes across the knife it is already pulling away from the OTC, so this is not an issue of the antiroll plate. I hadn't considered variations in temperature within the machine - I will check for that next time. I wondered if it would help if the knife was colder than the tissue - anyone tried this? A couple of pellets of dry ice on the knife holder perhaps? I will definitely try embedding the tissue in OTC before freezing next time, as well as infusing with OTC before freezing. But I wonder how to make a suitable mold? Someone has suggested aluminum foil. I ususally use re-useable plastic rings if the tissue is small enough, or else cut a piece of the bulb from a disposable plastic pasteur pipette for larger pieces, which I can just cut away after freezing. Also I understand from the comments here that mixing dry ice directly into the isopentane, rather than having the isopentance suspended above liquid nitrogen will give a more consistent temperature for snap freezing - another modification I will try next time. I had already settled on -26C for sectioning, as suggested by Gayle. I need to open up the intestine for my manipulations, so infusing the tube directly with OTC is not an option for me. But I need sections only 1 cm long, so it rolls up into a reasonably compact form. I have been rolling with villi out so they will be spread apart better, but I have also seen images rolled villi in - any preferences out there? So thanks everyone, hopefully my next attempts at freezing & sectioning will go much smoother. That brings me to my next problem, which I will describe in a separate post :) _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From ana.merino-trigo <@t> wanadoo.fr Mon Oct 3 13:07:28 2005 From: ana.merino-trigo <@t> wanadoo.fr (ana.merino-trigo) Date: Mon Oct 3 13:07:54 2005 Subject: [Histonet] FineFix fixative Message-ID: <009b01c5c845$51e751a0$02fea8c0@BABA> Hi everybody, Does anyone have any experience with FineFix fixative as a formalin substitute? We will run some test, however, I would really appreciate any comments. Thanks in advance, Ana (http://www.milestonemed.com/faq-finefix.php) What is FineFIX? FineFIX concentrate is a water-based additive. When added to ethanol, it creates a formalin-free fixative (the concentrate is not the actual fixative) for use in routine histology and molecular analysis. Its patented mix of chemical additives is reconstituted into a working solution by the addition of ethanol. How does FineFIX compare with Ethanol fixation? FineFIX is not subject to the disadvantages normally associated with Ethanol fixation (shrinkage, vacuolization, and pyknotic nuclei). Which tissue types has FineFIX been tested on, and for how long? The fixative has been in routine use for all specimen types in typical surgical pathology labs for continuous periods of up to two years. The quality of histology for tissues fixed in these labs shows absolutely the same appearance as that of the original samples. What is the effect of FineFIX on Epitope Retrieval and the subsequent Immunohistochemical technique? As FineFIX is not a cross-linking fixative, there is no need for aggressive epitope retrieval. It is recommended not to exceed a temperature of 100?C, for a duration of 5-15 minutes, during epitope retrieval procedures. The exact timing will depend on the type of antigen under evaluation and the immunohistochemical method performed by the laboratory. What are the benefits of FineFIX for molecular techniques? Improved recovery of DNA and RNA compared to formalin-fixed tissues. Thanks in advance, Ana Merino-Trigo, PhD Molecular Pathology - Genomic Sciencies Centre de Recherche de Paris, Magendie Sanofi-Aventis 13, Quai Jules Guesde 94403 Vitry sur Seine - France Telephone : +0033(0) 1589 38911 Fax : +0033(0) 1589 33344 E-mail : ana.merino-trigo@sanofi-aventis.com ana.merino-trigo@wanadoo.fr From LINDA.MARGRAF <@t> childrens.com Mon Oct 3 13:22:51 2005 From: LINDA.MARGRAF <@t> childrens.com (LINDA MARGRAF) Date: Mon Oct 3 13:23:29 2005 Subject: [Histonet] Microtome knives Message-ID: Dear Histonetters: I received an email from Cathy that says the following: I have available fourteen (14) - 160 mm microtome knives FREE to anyone who wants to pay for shipping. They can email me at cslarue@tylerhospital.com. Thanks, Cathy LaRue Please contact her directly if you are interested. I don't personally know her or know any details about the knives. Linda M Histonet administrator From Pat.Bell <@t> UCHSC.edu Mon Oct 3 13:51:33 2005 From: Pat.Bell <@t> UCHSC.edu (Pat.Bell@UCHSC.edu) Date: Mon Oct 3 13:51:50 2005 Subject: [Histonet] HER 3 Message-ID: <4762F4C84CD6144A91A7273EB0E999670D53FC@hscex1> Hi, Does anyone have a protocol for HER 3? Thanks, Pat Bell University of Co. Health Sciences Center Medical Oncology MS 8117 PO Box 6511 Aurora, Co 80045 303-724-0576 pat.bell@uchsc.edu From natalia_zinchenko <@t> hotmail.com Mon Oct 3 14:18:56 2005 From: natalia_zinchenko <@t> hotmail.com (Zinchenko Natalia) Date: Mon Oct 3 14:19:15 2005 Subject: [Histonet] live/dead protocol for liver Message-ID: Does anyone know where I can get Live/Dead protocol for liver? From pruegg <@t> ihctech.net Mon Oct 3 14:45:39 2005 From: pruegg <@t> ihctech.net (Patsy Ruegg) Date: Mon Oct 3 14:46:04 2005 Subject: [Histonet] RE: Tissue fixation help In-Reply-To: Message-ID: <200510031945.j93JjYAA009581@chip.viawest.net> I too have thawed tissue in rt formalin and then processed and embedded in paraffin which good morphology and IHC results (haven't tried with antibodies to phosphorylated sites but other IHC has been fine, it all depends on if the samples was properly frozen in the first place) I tried to do this with some heart tissue which had just been placed in a -80dc freezer with out snap freezing in OCT and it was awful, but the frozen section was bad as well. Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of C.M. van der Loos Sent: Sunday, October 02, 2005 11:43 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: Tissue fixation help Angela, Long time ago we thawed a frozen tissue sample directly in (room temp.) NBF (24-48 hs). Morphology was kept surprisingly well. Sorry, at that time phosphorylated antibodies were not even existant.... Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands phone: +31 20 5665631 Date: Fri, 30 Sep 2005 08:25:24 -0400 From: Angela McNabola Subject: [Histonet] Tissue fixation help To: [1]Histonet@lists.utsouthwestern.edu Good Morning, Does anybody on the histonet have any experience with taking tissue that has been snap frozen in minus 80, and then "thawed" out and formalin fixed and paraffin embedded? I won't get into the reasons why we have to do this, but does anyone have a protocol (or just slowly thaw it out and fix it??). How did your IHC turn out, especially when using phosphorlyated antibodies? Did you get good morphology of your tissue? thanks in advance! Angela Angela McNabola, MS HT(ASCP)SLS, QIHC Bayer Healthcare 400 Morgan Lane West Haven, CT 06516 203-812-5001 angela.mcnabola.b@bayer.com References 1. mailto:Histonet@lists.utsouthwestern.edu _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From pruegg <@t> ihctech.net Mon Oct 3 14:48:01 2005 From: pruegg <@t> ihctech.net (Patsy Ruegg) Date: Mon Oct 3 14:48:12 2005 Subject: [Histonet] immuno exam. In-Reply-To: Message-ID: <200510031947.j93JltAA010246@chip.viawest.net> If you join the NSH Immunohistochemistry Resource Group online at www.ihcrg.org you will have access to the ws material presented by Patsy Ruegg and Ethel Macrea at the NSH S/C in Florida. This material is now on the IHCRG website for members of the group. Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of KMB1904@aol.com Sent: Sunday, October 02, 2005 5:07 AM To: histonet@pathology.swmed.edu Subject: [Histonet] immuno exam. Does anyone know where I can get learning materials for the immuno exam for the ASCP?? Thanks _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From cwscouten <@t> myneurolab.com Mon Oct 3 14:53:35 2005 From: cwscouten <@t> myneurolab.com (Charles Scouten) Date: Mon Oct 3 14:54:31 2005 Subject: [Histonet] RE: Tissue fixation help Message-ID: <5784D843593D874C93E9BADCB87342AB9166D1@tpiserver03.Coretech-holdings.com> Just very curious. Why would anybody paraffin embed after the tissue had already been frozen? Either way gets it hard enough to cut, which is the purpose of both procedures. Cordially, Charles W. Scouten, Ph.D. myNeuroLab.com 5918 Evergreen Blvd. St. Louis, MO 63134 Ph: 314 522 0300 x 342 FAX 314 522 0377 cwscouten@myneurolab.com http://www.myneurolab.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Patsy Ruegg Sent: Monday, October 03, 2005 2:46 PM To: 'C.M. van der Loos'; Histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: Tissue fixation help I too have thawed tissue in rt formalin and then processed and embedded in paraffin which good morphology and IHC results (haven't tried with antibodies to phosphorylated sites but other IHC has been fine, it all depends on if the samples was properly frozen in the first place) I tried to do this with some heart tissue which had just been placed in a -80dc freezer with out snap freezing in OCT and it was awful, but the frozen section was bad as well. Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of C.M. van der Loos Sent: Sunday, October 02, 2005 11:43 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: Tissue fixation help Angela, Long time ago we thawed a frozen tissue sample directly in (room temp.) NBF (24-48 hs). Morphology was kept surprisingly well. Sorry, at that time phosphorylated antibodies were not even existant.... Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands phone: +31 20 5665631 Date: Fri, 30 Sep 2005 08:25:24 -0400 From: Angela McNabola Subject: [Histonet] Tissue fixation help To: [1]Histonet@lists.utsouthwestern.edu Good Morning, Does anybody on the histonet have any experience with taking tissue that has been snap frozen in minus 80, and then "thawed" out and formalin fixed and paraffin embedded? I won't get into the reasons why we have to do this, but does anyone have a protocol (or just slowly thaw it out and fix it??). How did your IHC turn out, especially when using phosphorlyated antibodies? Did you get good morphology of your tissue? thanks in advance! Angela Angela McNabola, MS HT(ASCP)SLS, QIHC Bayer Healthcare 400 Morgan Lane West Haven, CT 06516 203-812-5001 angela.mcnabola.b@bayer.com References 1. mailto:Histonet@lists.utsouthwestern.edu _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Barry.R.Rittman <@t> uth.tmc.edu Mon Oct 3 15:02:45 2005 From: Barry.R.Rittman <@t> uth.tmc.edu (Rittman, Barry R) Date: Mon Oct 3 15:03:01 2005 Subject: [Histonet] RE: Tissue fixation help Message-ID: Several reasons Perhaps the major one is that frozen and paraffin sections of equal thickness show a different morphology. Frozen tissue is said to have a general shrinkage of soft tissues of around 2-5% while in paraffin this may reach 25 -30%. Sections of equal thickness therefore contain different volumes of tissue and will stain to different degrees. The final image therefore does not always allow for easy comparison. Barry -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Charles Scouten Sent: Monday, October 03, 2005 2:54 PM To: Patsy Ruegg; C.M. van der Loos; Histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: Tissue fixation help Just very curious. Why would anybody paraffin embed after the tissue had already been frozen? Either way gets it hard enough to cut, which is the purpose of both procedures. Cordially, Charles W. Scouten, Ph.D. myNeuroLab.com 5918 Evergreen Blvd. St. Louis, MO 63134 Ph: 314 522 0300 x 342 FAX 314 522 0377 cwscouten@myneurolab.com http://www.myneurolab.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Patsy Ruegg Sent: Monday, October 03, 2005 2:46 PM To: 'C.M. van der Loos'; Histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: Tissue fixation help I too have thawed tissue in rt formalin and then processed and embedded in paraffin which good morphology and IHC results (haven't tried with antibodies to phosphorylated sites but other IHC has been fine, it all depends on if the samples was properly frozen in the first place) I tried to do this with some heart tissue which had just been placed in a -80dc freezer with out snap freezing in OCT and it was awful, but the frozen section was bad as well. Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of C.M. van der Loos Sent: Sunday, October 02, 2005 11:43 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: Tissue fixation help Angela, Long time ago we thawed a frozen tissue sample directly in (room temp.) NBF (24-48 hs). Morphology was kept surprisingly well. Sorry, at that time phosphorylated antibodies were not even existant.... Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands phone: +31 20 5665631 Date: Fri, 30 Sep 2005 08:25:24 -0400 From: Angela McNabola Subject: [Histonet] Tissue fixation help To: [1]Histonet@lists.utsouthwestern.edu Good Morning, Does anybody on the histonet have any experience with taking tissue that has been snap frozen in minus 80, and then "thawed" out and formalin fixed and paraffin embedded? I won't get into the reasons why we have to do this, but does anyone have a protocol (or just slowly thaw it out and fix it??). How did your IHC turn out, especially when using phosphorlyated antibodies? Did you get good morphology of your tissue? thanks in advance! Angela Angela McNabola, MS HT(ASCP)SLS, QIHC Bayer Healthcare 400 Morgan Lane West Haven, CT 06516 203-812-5001 angela.mcnabola.b@bayer.com References 1. mailto:Histonet@lists.utsouthwestern.edu _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rjbuesa <@t> yahoo.com Mon Oct 3 15:55:47 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Mon Oct 3 15:56:04 2005 Subject: [Histonet] RE: Tissue fixation help In-Reply-To: Message-ID: <20051003205547.20871.qmail@web61211.mail.yahoo.com> Sometimes the pathologists give a diagnosis on a frozen section "pending" of a definitive diagnosis when "permanent sections" (i.e. paraffin sections are done). In our laboratory it was routine to process what was left of the tissue used for the frozen section and one of the elements to take into consideration was the size and shape of the paraffin sections vs. that of the frozen section. In our laboratory we routinely obtained a reduction in size of the order of 20 to 25% (sometimes a little more or a little less, depending on the nature of the tissue: the harder it was, the less shrinkage). Rene J. "Rittman, Barry R" wrote: Several reasons Perhaps the major one is that frozen and paraffin sections of equal thickness show a different morphology. Frozen tissue is said to have a general shrinkage of soft tissues of around 2-5% while in paraffin this may reach 25 -30%. Sections of equal thickness therefore contain different volumes of tissue and will stain to different degrees. The final image therefore does not always allow for easy comparison. Barry -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Charles Scouten Sent: Monday, October 03, 2005 2:54 PM To: Patsy Ruegg; C.M. van der Loos; Histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: Tissue fixation help Just very curious. Why would anybody paraffin embed after the tissue had already been frozen? Either way gets it hard enough to cut, which is the purpose of both procedures. Cordially, Charles W. Scouten, Ph.D. myNeuroLab.com 5918 Evergreen Blvd. St. Louis, MO 63134 Ph: 314 522 0300 x 342 FAX 314 522 0377 cwscouten@myneurolab.com http://www.myneurolab.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Patsy Ruegg Sent: Monday, October 03, 2005 2:46 PM To: 'C.M. van der Loos'; Histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: Tissue fixation help I too have thawed tissue in rt formalin and then processed and embedded in paraffin which good morphology and IHC results (haven't tried with antibodies to phosphorylated sites but other IHC has been fine, it all depends on if the samples was properly frozen in the first place) I tried to do this with some heart tissue which had just been placed in a -80dc freezer with out snap freezing in OCT and it was awful, but the frozen section was bad as well. Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of C.M. van der Loos Sent: Sunday, October 02, 2005 11:43 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: Tissue fixation help Angela, Long time ago we thawed a frozen tissue sample directly in (room temp.) NBF (24-48 hs). Morphology was kept surprisingly well. Sorry, at that time phosphorylated antibodies were not even existant.... Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands phone: +31 20 5665631 Date: Fri, 30 Sep 2005 08:25:24 -0400 From: Angela McNabola Subject: [Histonet] Tissue fixation help To: [1]Histonet@lists.utsouthwestern.edu Good Morning, Does anybody on the histonet have any experience with taking tissue that has been snap frozen in minus 80, and then "thawed" out and formalin fixed and paraffin embedded? I won't get into the reasons why we have to do this, but does anyone have a protocol (or just slowly thaw it out and fix it??). How did your IHC turn out, especially when using phosphorlyated antibodies? Did you get good morphology of your tissue? thanks in advance! Angela Angela McNabola, MS HT(ASCP)SLS, QIHC Bayer Healthcare 400 Morgan Lane West Haven, CT 06516 203-812-5001 angela.mcnabola.b@bayer.com References 1. mailto:Histonet@lists.utsouthwestern.edu _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From david.kinsley <@t> spcorp.com Mon Oct 3 16:17:48 2005 From: david.kinsley <@t> spcorp.com (Kinsley, David) Date: Mon Oct 3 16:18:15 2005 Subject: [Histonet] Whole Rat Paw Processing Message-ID: <79A75F7AD4BFAE4CA27C8AABE3F75D9A0E6B65@KENMSG22.us.schp.com> Hi Histonetters, I'm going to be working on a project where I will need to collect, process, and cut whole rat paws from the ankle to the toes. Does anyone have any experience with these? I'm in need of larger size processing cassettes and embedding molds, decalcification times/processing times, as well as information on chuck adapters for the microtome. Any advice/information is greatly appreciated. David Kinsley Schering-Plough Research Institute Kenilworth, NJ 07033 908-740-4669 ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. From JUDIEWHIT <@t> aol.com Mon Oct 3 18:11:54 2005 From: JUDIEWHIT <@t> aol.com (JUDIEWHIT@aol.com) Date: Mon Oct 3 18:12:15 2005 Subject: [Histonet] Re: Help Message-ID: <7f.67c1d32e.307314ba@aol.com> TO All Histotechs: Please send me any and all references/addresses for any articles on Histologists, Microtomy,Chronic Repetitive Trauma, and Techniques to Avoid Injury. I need reprints from any articles or studies with sources on the above. Thank you, Judie Whitcomb HT/HTL(ASCP) From ihctech2000 <@t> yahoo.com Mon Oct 3 18:35:32 2005 From: ihctech2000 <@t> yahoo.com (Sun Zhon) Date: Mon Oct 3 18:35:54 2005 Subject: [Histonet] Elution buffer on frozen tissues Message-ID: <20051003233532.56026.qmail@web53713.mail.yahoo.com> Hi All, I am going to run a dual chromogen staining on frozen tissues. I am wondering whether I can use the elution buffer to elute the 1st antibody complex? Will the elution buffer destroy morphology of the frozen tissues? Thanks in advance for the help. --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From cfavara <@t> niaid.nih.gov Mon Oct 3 19:10:05 2005 From: cfavara <@t> niaid.nih.gov (Favara, Cynthia (NIH/NIAID)) Date: Mon Oct 3 19:10:35 2005 Subject: [Histonet] RE: Tissue fixation help Message-ID: Oh you know sometimes the PI is not thinking clearly or at all and thinks that frozen fixed whatever can be used for anything and the animal model takes about 2 years for development of disease. Then they want everything and don't want to work with frozen sections it goes on and on ......... Need to remember in the clinical world sometimes only a small biopsy and need to confirm with FFPE sections - the gold standard! Just a few - sorry for the snippy attitude it has been a day - c Cynthia Favara NIAID/NIH/RML/LPVD 903 South 4th Street Hamilton, MT 59840 406-363-9317 Disclaimer: The information in this e-mail and any of its attachments is confidential and may contain sensitive information. It should not be used by anyone who is not the original intended recipient. If you have received this e-mail in error please inform the sender and delete it from your mailbox or any other storage devices. National Institute of Allergy and Infectious Diseases shall not accept liability for any statements made that are sender's own and not expressly made on behalf of the NIAID by one of its representatives -----Original Message----- From: Charles Scouten [mailto:cwscouten@myneurolab.com] Sent: Monday, October 03, 2005 12:54 PM To: Patsy Ruegg; C.M. van der Loos; Histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: Tissue fixation help Just very curious. Why would anybody paraffin embed after the tissue had already been frozen? Either way gets it hard enough to cut, which is the purpose of both procedures. Cordially, Charles W. Scouten, Ph.D. myNeuroLab.com 5918 Evergreen Blvd. St. Louis, MO 63134 Ph: 314 522 0300 x 342 FAX 314 522 0377 cwscouten@myneurolab.com http://www.myneurolab.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Patsy Ruegg Sent: Monday, October 03, 2005 2:46 PM To: 'C.M. van der Loos'; Histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: Tissue fixation help I too have thawed tissue in rt formalin and then processed and embedded in paraffin which good morphology and IHC results (haven't tried with antibodies to phosphorylated sites but other IHC has been fine, it all depends on if the samples was properly frozen in the first place) I tried to do this with some heart tissue which had just been placed in a -80dc freezer with out snap freezing in OCT and it was awful, but the frozen section was bad as well. Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of C.M. van der Loos Sent: Sunday, October 02, 2005 11:43 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: Tissue fixation help Angela, Long time ago we thawed a frozen tissue sample directly in (room temp.) NBF (24-48 hs). Morphology was kept surprisingly well. Sorry, at that time phosphorylated antibodies were not even existant.... Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands phone: +31 20 5665631 Date: Fri, 30 Sep 2005 08:25:24 -0400 From: Angela McNabola Subject: [Histonet] Tissue fixation help To: [1]Histonet@lists.utsouthwestern.edu Good Morning, Does anybody on the histonet have any experience with taking tissue that has been snap frozen in minus 80, and then "thawed" out and formalin fixed and paraffin embedded? I won't get into the reasons why we have to do this, but does anyone have a protocol (or just slowly thaw it out and fix it??). How did your IHC turn out, especially when using phosphorlyated antibodies? Did you get good morphology of your tissue? thanks in advance! Angela Angela McNabola, MS HT(ASCP)SLS, QIHC Bayer Healthcare 400 Morgan Lane West Haven, CT 06516 203-812-5001 angela.mcnabola.b@bayer.com References 1. mailto:Histonet@lists.utsouthwestern.edu _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From petepath <@t> yahoo.com Mon Oct 3 20:39:59 2005 From: petepath <@t> yahoo.com (Stephen Peters M.D.) Date: Mon Oct 3 20:40:29 2005 Subject: [Histonet] RE: Tissue fixation help Message-ID: <20051004014000.2682.qmail@web30403.mail.mud.yahoo.com> Pretty much all of the frozen sections done in hospital pathology departments are then thawed and paraffin embedded. Some call this the frozen section control. I have some a few pics on my web site that show freeze artifact next to the frozen controls. About midway down this page. http://pathologyinnovations.com/new_page_3.htm The nuclear chromatin is a bit less defined and overall have a slightly smudged quality compared to tissue the not previously frozen. The actual ice crystal artifact seems to dissipate after processing the tissue. As far as immunostains go, I have gotten satisfactory results with previously frozen samples with maybe a bit more background. The finicky ones are probably more finicky. How do others feel about immunos on frozen controls? Stephen From jmaass <@t> info2000.net Tue Oct 4 00:01:33 2005 From: jmaass <@t> info2000.net (Janet Maass) Date: Tue Oct 4 00:06:30 2005 Subject: [Histonet] replacement chemical Message-ID: <000e01c5c8a0$b1d25000$ab2ae404@oemcomputer> Does anyone know of a chemical replacement for 2,3,5-Triphenyltetrazolium chloride? This chemical has not been available for the past few months and may not be available for a few more months depending on the testing results. The government is considering a reclassification depending on the testing results so currently no one can purchase it. I have not had good results using nitro blue tetrazolium as a replacement. I am using it as an oxidation and reduction substance to identify on fresh tissue the area where damaged tissue ends and undamaged tissue begins in research. Should someone have a procedure I would appreciate it. In my searches on Pubmed I find articles using nitro blue tetrazolium on bacteria and the likes, but not fresh tissue. Thanks in advance, Janet Maass From jkiernan <@t> uwo.ca Tue Oct 4 00:18:38 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Tue Oct 4 00:19:02 2005 Subject: [Histonet] Re: Help References: <7f.67c1d32e.307314ba@aol.com> Message-ID: <434210AE.87BBB4BD@uwo.ca> Dear Judie Whitcomb, Who are you, and why should we all put in several unpaid hours to do your work for you? Read what you wrote (cited below), and think about what you are requesting! You sign as an HT/HTL(ASCP), indicating that in the USA you have passed public exams in most of the subjects you ask about. John Kiernan, London, Canada. ------------------------------ JUDIEWHIT@aol.com wrote: > > TO All Histotechs: > Please send me any and all references/addresses for any articles on > Histologists, > Microtomy,Chronic Repetitive Trauma, and Techniques to Avoid Injury. > I need reprints from any articles or studies with sources on the above. > Thank you, > Judie Whitcomb HT/HTL(ASCP) From lpwenk <@t> sbcglobal.net Tue Oct 4 04:12:55 2005 From: lpwenk <@t> sbcglobal.net (Lee & Peggy Wenk) Date: Tue Oct 4 04:13:38 2005 Subject: [Histonet] immuno exam. In-Reply-To: Message-ID: <1083705854-404468982@pathology.swmed.edu> 1. Dako has their IHC staining book (3rd edition) and an IHC glossary available for download. Click under "literature" on left side http://www.dakocytomation.us/index/us_support_literature_index/ 2. LabVision has several free IHC on-line tutorials. Need to register first, but can earn NSH contact hours (continuing education). http://www.labvision.com/indexTutorial.cfm 3. NSH has a self-assessment booklet (#5) on IHC, ISH, EM, etc., about 100 multiple choice questions. $15 for NSH members, $35 for non-members. http://www.nsh.org/education/materials.html 4. NSH has teleconferences coming up on IHC. Dec. 21 & Jan 18 are basic IHC part I and II. The rest of 2006 is not yet on line, but I think the February teleconference is on IHC of cytokeratins. And then later in 2006, there is one on the lab math of titering/dilutions. They are $90 each ($100 if get a tape afterwards that everyone in the lab can listen to during some free time later). Either way, can earn 1 contact hour continuing education from NSH for each person in lab. http://www.nsh.org/education/teleconf2002.html Hope that helps. Peggy A. Wenk, HTL(ASCP)SLS William Beaumont Hospital Royal Oak, MI 48073 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of KMB1904@aol.com Sent: Sunday, October 02, 2005 8:07 AM To: histonet@pathology.swmed.edu Subject: [Histonet] immuno exam. Does anyone know where I can get learning materials for the immuno exam for the ASCP?? Thanks _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From LJApuzzio <@t> msn.com Tue Oct 4 06:04:35 2005 From: LJApuzzio <@t> msn.com (Louis Apuzzio) Date: Tue Oct 4 06:05:01 2005 Subject: [Histonet] Employment Message-ID: I am looking for employment in Histology. I am HT eligible and also CT(ASCP) Able to do cutting,embedding,frozen sectioning,grossing,special stains and autopsies. Very dependable and consistent. Availability is immediate Louis J. Apuzzio From rjbuesa <@t> yahoo.com Tue Oct 4 07:31:16 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Tue Oct 4 07:31:32 2005 Subject: [Histonet] Re: Help In-Reply-To: <434210AE.87BBB4BD@uwo.ca> Message-ID: <20051004123116.42978.qmail@web61222.mail.yahoo.com> Well said John! John Kiernan wrote:Dear Judie Whitcomb, Who are you, and why should we all put in several unpaid hours to do your work for you? Read what you wrote (cited below), and think about what you are requesting! You sign as an HT/HTL(ASCP), indicating that in the USA you have passed public exams in most of the subjects you ask about. John Kiernan, London, Canada. ------------------------------ JUDIEWHIT@aol.com wrote: > > TO All Histotechs: > Please send me any and all references/addresses for any articles on > Histologists, > Microtomy,Chronic Repetitive Trauma, and Techniques to Avoid Injury. > I need reprints from any articles or studies with sources on the above. > Thank you, > Judie Whitcomb HT/HTL(ASCP) _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From liz <@t> premierlab.com Tue Oct 4 08:40:04 2005 From: liz <@t> premierlab.com (Elizabeth Chlipala) Date: Tue Oct 4 08:40:47 2005 Subject: [Histonet] Whole Rat Paw Processing In-Reply-To: <79A75F7AD4BFAE4CA27C8AABE3F75D9A0E6B65@KENMSG22.us.schp.com> Message-ID: <000001c5c8e9$21958770$a7d48a80@AMY> David Is there a reason why you need to process the specimens whole. I have processed both rat ankles and rear paws, but separately, the ankle being sectioned to demonstrate the tibia and tibial tarsal joints and associated inflammation (arthritis model) and the paws to demonstrate the digits. E-mail me back with some more specifics and I can probably help you out, but I need a bit more information. On our web page there are a few images of some rat ankles. And if you look at the white papers section of the web page there is a poster that covers ankle preparation. I have other information also that will send off the histonet. If you do need to process the rear ankle and paw together you might be able to just use mega cassettes and the largest base molds, the specimens might fit, if that's not big enough I would either use surgipaths system for the larger cassettes if that's not a possibility the old steel L's should work with a microtome chuck that will support those blocks. Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC P.O. Box 18592 Boulder, Colorado 80308 Office: (303) 735-5001 Fax: (303) 735-3540 liz@premierlab.com www.premierlab.com Ship to Address: Premier Laboratory University of Colorado MCDB, Room A3B40 Boulder, Colorado 80309 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Kinsley, David Sent: Monday, October 03, 2005 2:18 PM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] Whole Rat Paw Processing Hi Histonetters, I'm going to be working on a project where I will need to collect, process, and cut whole rat paws from the ankle to the toes. Does anyone have any experience with these? I'm in need of larger size processing cassettes and embedding molds, decalcification times/processing times, as well as information on chuck adapters for the microtome. Any advice/information is greatly appreciated. David Kinsley Schering-Plough Research Institute Kenilworth, NJ 07033 908-740-4669 ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Janet.Bonner <@t> FLHOSP.ORG Tue Oct 4 11:05:12 2005 From: Janet.Bonner <@t> FLHOSP.ORG (Bonner, Janet) Date: Tue Oct 4 11:05:54 2005 Subject: [Histonet] removing silver from counters, etc. Message-ID: <07AB60D5D7B9754EBF56F360F98D083DEB4409@fh2k093.fhmis.net> Tarn-x works wonders. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Hofecker, Jennifer L Sent: Friday, September 23, 2005 9:24 AM To: histonet Subject: [Histonet] removing silver from counters, etc. Happy Friday everyone. Can somebody remind me what solutions will remove silver nitrate from cabinets, floors, counters, etc.? Bleach just changes these spots to different shades of brown. I remember using solution from hair perms in the past but I am sure that there are other reliable ways. Thanks for any advice. Our lab is starting to resemble a dalmation! You know how silver solution likes to migrate from the coplin jars! Thoughts and prayers go out to Gulf Coast residents. Jennifer Jennifer Hofecker, HT (ASCP) Vanderbilt University Medical Center Division of Neuropathology (615) 343-0083 (615) 343-7089 fax _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rjbuesa <@t> yahoo.com Tue Oct 4 11:07:54 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Tue Oct 4 11:08:14 2005 Subject: [Histonet] Update on Tricks if the Trade (TOTs). Message-ID: <20051004160754.92338.qmail@web61214.mail.yahoo.com> To my fellow histotechnologists: As I promised on 9/20/05 when I posted the request for TOTs to be edited/published in the Journal of Histotechnology, the following is a summary of the TOTs received: 1- how to cool paraffin blocks by using the denser than air CO2 gas coming from a funnel placed above the block. 2- add eosin in the first absolute alcohol to stain small transparent specimens. 3- place "roght cut" blocks on very wet 4X4 gauze pieces for a few minutes before cutting the final sections. 4- for cell proliferation studies in rodents always simultaneously process a piece of ileum or jejunum as control. 5-test the DAB with a few microlitres of peroxidase labelled Ab to determine if DAB is OK. 6- soak faced blocks in a small container with crushed ice, some water and a few drops of amonia, before cutting final sections. And, unfortunately, that is all!!! No more TOTs were received! I have been really disapointed because from all the histotechnologists using Histonet, after reading so many queries and answers, I know there is wealth of TOTs out there. I realize that it is easier to just give some quick advise to some "histoneter" in need of help, than to squeeze one's brain to come with something we every day do as a TOT. At this moment I am preparing to edit TOTs dealing "toe nails" (an abundant subject of discussion in Histonet since 2001); this and any other important issue is going to help all of us, but specially those new to the field. This is part of the mission of the Journal of Histotechnology and we all with more experience have the moral duty of helping those less experienced. I only hope that all the "Histoneters" will contribute with their TOTs for the benefit of all. Rene J. Buesa --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From lrichey <@t> u.washington.edu Tue Oct 4 11:48:20 2005 From: lrichey <@t> u.washington.edu (Lori Richey) Date: Tue Oct 4 11:48:41 2005 Subject: [Histonet] RE: Tissue fixation help In-Reply-To: <5784D843593D874C93E9BADCB87342AB9166D1@tpiserver03.Coretech-holdings.com> References: <5784D843593D874C93E9BADCB87342AB9166D1@tpiserver03.Coretech-holdings.com> Message-ID: <4342B254.4020405@u.washington.edu> It's a common protocol to embed tissue in paraffiin after its been reviewed as a frozen section. The tissue is frozen for quick diagnosis while the patient is still in surgery. It is then melted down and submitted for paraffin sections. Charles Scouten wrote: >Just very curious. Why would anybody paraffin embed after the tissue >had already been frozen? Either way gets it hard enough to cut, which is >the purpose of both procedures. > > >Cordially, >Charles W. Scouten, Ph.D. >myNeuroLab.com >5918 Evergreen Blvd. >St. Louis, MO 63134 >Ph: 314 522 0300 x 342 >FAX 314 522 0377 >cwscouten@myneurolab.com >http://www.myneurolab.com > > >-----Original Message----- >From: histonet-bounces@lists.utsouthwestern.edu >[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Patsy >Ruegg >Sent: Monday, October 03, 2005 2:46 PM >To: 'C.M. van der Loos'; Histonet@lists.utsouthwestern.edu >Subject: RE: [Histonet] RE: Tissue fixation help > >I too have thawed tissue in rt formalin and then processed and embedded >in paraffin which good morphology and IHC results (haven't tried with >antibodies to phosphorylated sites but other IHC has been fine, it all >depends on if the samples was properly frozen in the first place) I >tried to do this with some heart tissue which had just been placed in a >-80dc freezer with out snap freezing in OCT and it was awful, but the >frozen section was bad as well. >Patsy > > >Patsy Ruegg, HT(ASCP)QIHC >IHCtech, LLC >Fitzsimmons BioScience Park >12635 Montview Blvd. Suite 216 >Aurora, CO 80010 >P-720-859-4060 >F-720-859-4110 >wk email pruegg@ihctech.net >web site www.ihctech.net > > >This email is confidential and intended solely for the use of the >Person(s) ('the intended recipient') to whom it was addressed. Any views >or opinions presented are solely those of the author. It may contain >information that is privileged & confidential within the meaning of >applicable law. Accordingly any dissemination, distribution, copying, or >other use of this message, or any of its contents, by any person other >than the intended recipient may constitute a breach of civil or criminal >law and is strictly prohibited. If you are NOT the intended recipient >please contact the sender and dispose of this e-mail as soon as >possible. > > >-----Original Message----- >From: histonet-bounces@lists.utsouthwestern.edu >[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of C.M. van >der Loos >Sent: Sunday, October 02, 2005 11:43 PM >To: Histonet@lists.utsouthwestern.edu >Subject: [Histonet] RE: Tissue fixation help > > > Angela, > > Long time ago we thawed a frozen tissue sample directly in >(room > temp.) NBF (24-48 hs). Morphology was kept surprisingly well. >Sorry, > at that time phosphorylated antibodies were not even existant.... > > Chris van der Loos, PhD > Dept. of Pathology > Academic Medical Center M2-230 > Meibergdreef 9 > NL-1105 AZ Amsterdam > The Netherlands > > phone: +31 20 5665631 > > Date: Fri, 30 Sep 2005 08:25:24 -0400 > From: Angela McNabola > Subject: [Histonet] Tissue fixation help > To: [1]Histonet@lists.utsouthwestern.edu > Good Morning, > Does anybody on the histonet have any experience with taking >tissue > that has > been snap frozen in minus 80, and then "thawed" out and formalin >fixed > and > paraffin embedded? > I won't get into the reasons why we have to do this, but does >anyone > have a > protocol (or just slowly thaw it out and fix it??). How did your >IHC > turn out, > especially when using phosphorlyated antibodies? Did you get >good > morphology of > your tissue? > thanks in advance! > Angela > Angela McNabola, MS HT(ASCP)SLS, QIHC > Bayer Healthcare > 400 Morgan Lane > West Haven, CT 06516 > 203-812-5001 > angela.mcnabola.b@bayer.com > >References > > 1. mailto:Histonet@lists.utsouthwestern.edu >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > From Tbarnhart <@t> primecare.org Tue Oct 4 11:55:53 2005 From: Tbarnhart <@t> primecare.org (Barnhart, Tammy) Date: Tue Oct 4 11:57:40 2005 Subject: [Histonet] AFB control Message-ID: <1779904B5E82D511914C00D0B793339205BFDA35@exchangent> I am getting very low on AFB control and wonder if anyone has extra and would be willing to trade? Please contact me directly and lets see if we can work something out. I have lots of odd controls and tissues I would be willing to trade. Tammy Barnhart Confidentiality Notice:This e-mail message is for sole use of intended recipient(s) and may contain confidential and privileged information. Any unauthorized review, use, disclosure, distribution, or copying is prohibited. If you are not the intended recipient, please contact the sender by replying to this e-mail and destroy/delete all copies of this e-mail message. From brett_connolly <@t> merck.com Tue Oct 4 12:25:30 2005 From: brett_connolly <@t> merck.com (Connolly, Brett M) Date: Tue Oct 4 12:26:20 2005 Subject: [Histonet] Racks for large slides Message-ID: <355C35514FEAC9458F75947F5270974D079638@usctmx1103.merck.com> We are embarking on some studies where we will be mounting large tissue sections on 5"x 7" slides. We got the slides easy enough at Brain Research Labs, but they don't offer any type of rack (or staining dishes) to go with the slides. I didn't see anything at MyNeuroLab either. We might be able to have some racks fabricated in-house, but it would take quite a while here. Any suggestions? Thanks, Brett Brett M. Connolly, Ph.D. Merck & Co., Inc. MRL, Imaging Research WP-44K PO Box 4 West Point, PA 19486 PH 215-652-2501 fax. 215-993-6803 e-mail. brett_connolly@merck.com ------------------------------------------------------------------------------ Notice: This e-mail message, together with any attachments, contains information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates (which may be known outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD and in Japan, as Banyu) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system. ------------------------------------------------------------------------------ From pmarcum <@t> vet.upenn.edu Tue Oct 4 12:32:07 2005 From: pmarcum <@t> vet.upenn.edu (Pamela Marcum) Date: Tue Oct 4 12:32:35 2005 Subject: [Histonet] Region II Meeting in King of Prussia PA Message-ID: <6.1.1.1.2.20051004132249.019d9380@mail.vet.upenn.edu> Good Afternoon to All, We have clinical, research and veterinary talks as well as a special CSI program day for Saturday!!! Region II (NSH) comprised of Delaware, New Jersey, Maryland and Pennsylvania (Including Washington DC, Virginia, and West Virginia without current active societies) will be hosting a meeting in King of Prussia PA on November 4th and 5th, 2005 at the Hilton Valley Forge, 251 DeKalb Pike (Route 202). The meeting program is available for any one wishing to come or for your review. Please e-mail me for your copy of the program TODAY and plan to join us for a great meeting. Best Regards, Pamela A Marcum Manager, Histology Special Procedures University of Pennsylvania School of Veterinary Medicine R.S. Reynolds Jr. CORL New Bolton Center 382 West Street Road Kennett Square, PA 19348 Phone - 610-925-6278 Fax - 610-925-8120 E-mail - pmarcum@vet.upenn.edu From angela.mcnabola.b <@t> bayer.com Tue Oct 4 13:53:23 2005 From: angela.mcnabola.b <@t> bayer.com (Angela McNabola) Date: Tue Oct 4 13:53:19 2005 Subject: [Histonet] Re: Tissue fixation help Message-ID: Hi all, I just wanted to thank everyone for your responses regarding fixation of the frozen tissues into formalin! I'll let you know how the staining turns out. Angela Angela McNabola MS, HT(ASCP)SLS, QIHC Scientist Bayer Helathcare 400 Morgan Lane West Haven, CT 06516 203-812-5001 angela.mcnabola.b@bayer.com From contact <@t> excaliburpathology.com Tue Oct 4 13:58:37 2005 From: contact <@t> excaliburpathology.com (Paula Pierce) Date: Tue Oct 4 13:58:52 2005 Subject: [Histonet] Large staining racks Message-ID: <20051004185838.19111.qmail@web50314.mail.yahoo.com> Try a photographic supply. We used racks for TEM & SEM negatives that were 3X5". Something like that should work. Paula Pierce, HTL(ASCP)HT Excalibur Pathology, Inc. 630 N. Broadway Moore, OK 73160 405-759-3953 contact@excaliburpathology.com www.excaliburpathology.com From mward <@t> wfubmc.edu Tue Oct 4 14:15:12 2005 From: mward <@t> wfubmc.edu (Martha Ward) Date: Tue Oct 4 14:16:35 2005 Subject: [Histonet] rocky mountain spotted fever antibody Message-ID: <61135F0455D33347B5AAE209B903A3041023A525@EXCHVS2.medctr.ad.wfubmc.edu> I am searching for a new vendor for our FITC-labeled R. rickettsii antibody. We used to get it from the CDC but according to my sources they are no longer providing the antibody. Does anyone out there know of another source for this antibody? Thanks in advance for your help. Martha Ward Wake Forest University Baptist Medical Center From alex <@t> fronine.com.au Tue Oct 4 21:19:28 2005 From: alex <@t> fronine.com.au (Alex Cross) Date: Tue Oct 4 21:21:50 2005 Subject: [Histonet] Carey Blair fixative Message-ID: <20051005022130.FTZP5164.omta01sl.mx.bigpond.com@NEWLABPC> Please help. I am after some information on Carey Blair Fixative. I would like to prepare some. Thanks in advance Alexander Cross Fronine Laboratories Australia From c.m.vanderloos <@t> amc.uva.nl Wed Oct 5 01:51:48 2005 From: c.m.vanderloos <@t> amc.uva.nl (C.M. van der Loos) Date: Wed Oct 5 01:52:10 2005 Subject: [Histonet] RE: Elution buffer on frozen tissues Message-ID: <1141dcd11441e3.11441e31141dcd@amc.uva.nl> Dear Sun Zhon, You better don't use an elution buffer on cryo's during sequential double staining at all. In fact you don't need it and with high affinity primaries it's not even effective. When performing sequential doublestaining one MUST apply DAB as chromogen after the first staining sequence. DAB is the only enzymatic reaction product that has the unique characteristic of sheltering the first set of immunoreagents effectively. This DAB-layer prevents cross-reaction with the second staining sequence. There is one important thing you should take care of: an appropriate dilution of the first step primary antibody. If this is too concentrated, the DAB sheltering effect is not completely effective and cross-reaction with the second staining sequence will occur. Good luck! Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands Date: Mon, 3 Oct 2005 16:35:32 -0700 (PDT) From: Sun Zhon Subject: [Histonet] Elution buffer on frozen tissues To: histonet@lists.utsouthwestern.edu Hi All, I am going to run a dual chromogen staining on frozen tissues. I am wondering whether I can use the elution buffer to elute the 1st antibody complex? Will the elution buffer destroy morphology of the frozen tissues? Thanks in advance for the help. From hmorrow <@t> cogeco.ca Wed Oct 5 06:20:31 2005 From: hmorrow <@t> cogeco.ca (Marg & Vern Morrow) Date: Wed Oct 5 06:20:36 2005 Subject: [Histonet] pepsin A Message-ID: <000601c5c99e$cc15e590$305fe218@your4o5qm9xfjg> We have pepsin A that was bought from BDH and is no longer available. Does anybody have another source? Tx Marg From SARAH.REEVES <@t> ekht.nhs.uk Wed Oct 5 06:36:16 2005 From: SARAH.REEVES <@t> ekht.nhs.uk (SARAH REEVES) Date: Wed Oct 5 07:29:21 2005 Subject: [Histonet] BSc PROJECT IDEAS Message-ID: Hi We have a student doing her BSc Biomedical Science who is looking for ideas for her final year project. She wants to specialise in Cellular Pathology although she does work mainly in Histology with an interest in special stains. We have just taken receipt of a TP 3000 so liquid based cytology is also an option. Any ideas please help! Cheers Sarah Reeves ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. Any views or opinions presented are solely those of the author and do not necessarily represent those of East Kent Hospitals NHS Trust. If you are not the intended recipient, be advised that you have received this email in error and that any use, dissemination, forwarding, printing or copying of this email is strictly prohibited. If you have received this email in error please notify the system manager at the following email address: root.postmaster@ekht.nhs.uk www.kentandmedway.nhs.uk This footnote also confirms that although this email message has been swept by MIMEsweeper for the presence of computer viruses, it is strongly recommended that you carry out your own virus scan of this message and any attachments. www.mimesweeper.com ********************************************************************** From rjbuesa <@t> yahoo.com Wed Oct 5 10:49:15 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Wed Oct 5 10:49:36 2005 Subject: [Histonet] Carey Blair fixative In-Reply-To: <20051005022130.FTZP5164.omta01sl.mx.bigpond.com@NEWLABPC> Message-ID: <20051005154915.44945.qmail@web61216.mail.yahoo.com> Hi Alex: The closest I can recognize is the Blair and Davies procedure for nerves in heart. If this is the subject of your study please let me know so I can send you the procedure. Rene J. Alex Cross wrote: Please help. I am after some information on Carey Blair Fixative. I would like to prepare some. Thanks in advance Alexander Cross Fronine Laboratories Australia _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From dobbin <@t> upei.ca Wed Oct 5 11:58:18 2005 From: dobbin <@t> upei.ca (Greg Dobbin) Date: Wed Oct 5 11:01:56 2005 Subject: [Histonet] Carey Blair fixative In-Reply-To: <20051005022130.FTZP5164.omta01sl.mx.bigpond.com@NEWLABPC> Message-ID: <4343CDEA.26938.354BF4@acad1.cs.upei.ca> Hi Alex, The only Carey-Blair reagent that I am familiar with is Carey-Blair transport medium for transport of microbiological specimens (usually swabs). A special formulation to be optimized for isolation of enteric organisms. Greg From: "Alex Cross" To: Date sent: Wed, 5 Oct 2005 12:19:28 +1000 Subject: [Histonet] Carey Blair fixative > Please help. I am after some information on Carey Blair Fixative. I > would like to prepare some. > > Thanks in advance Alexander Cross Fronine Laboratories > Australia > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From ADeng <@t> som.umaryland.edu Wed Oct 5 11:03:29 2005 From: ADeng <@t> som.umaryland.edu (Deng, April) Date: Wed Oct 5 11:03:44 2005 Subject: [Histonet] Histotech job Message-ID: Hi, Every one: There is a job opening for histolab in Dermatology Department, University of Maryland in Baltimore. Is anybody interested? April Deng <> From ADeng <@t> som.umaryland.edu Wed Oct 5 11:54:10 2005 From: ADeng <@t> som.umaryland.edu (Deng, April) Date: Wed Oct 5 11:54:23 2005 Subject: [Histonet] Histotechnician Job Opportunity Message-ID: The Dermatology Department, University of Maryland in Baltimore has an immediate opening for a technician in our histology laboratory. The main task of the job is to prepare high quality stained sections of patient skin tissues for microscopic examination by Pathologist. The specimen number is limited and the working hours are flexible. Education/Training/Certification/Licensure: AA/AS. Minimum of 2-3 years full-time experience in histopathology working in a clinical and/or research setting under the direction of a board certified pathologist. General knowledge and experience of performing histochemical-based staining procedures (e.g., H&E) in is required. Need to have the ability to work independently and good organizational skills with attention to detail. ASCP histotechnician certification is desired. Salary is based on experience and an excellent benefits package is offered. Applicants should submit a CV, names of three (3) references and salary history to: April Deng, M.D., Ph.D. Director of the Dermatopathology laboratory, University of Maryland, Department of Dermatology, 405 W. Redwood Street, 6th FL, Baltimore, MD 21201 TEL 410-328-5766 FAX 410-328-0098, E-mail: adeng@som.umaryland.edu From pruegg <@t> ihctech.net Wed Oct 5 15:37:50 2005 From: pruegg <@t> ihctech.net (Patsy Ruegg) Date: Wed Oct 5 15:38:07 2005 Subject: [Histonet] Fetal mouse samples Message-ID: <200510052037.j95Kbf9O022163@chip.viawest.net> I am processing whole fetal mouse samples (these are pretty big, they have skin developed) and the investigator wants to have sections so that both lungs show up at the same time in the same section. Can someone recommend to me a way to dissect these to open them up so that they will fix and process well but still maintain the whole body architecture for sectioning. Thank you, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. From rjbuesa <@t> yahoo.com Wed Oct 5 16:00:18 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Wed Oct 5 16:00:38 2005 Subject: [Histonet] Fetal mouse samples In-Reply-To: <200510052037.j95Kbf9O022163@chip.viawest.net> Message-ID: <20051005210019.5811.qmail@web61211.mail.yahoo.com> Patsy: Many years ago I had to do that. Of the three "classical" sectioning planes (transverse, sagittal and frontal) the one to use is the frontal. Rest the embryo flat on its back and cut it open starting at nose/mouth level all the way back to the vertebral column in a way that the back half of the embryo can be separated from the front half. Try to harden it first with formaldehyde (4-6 hours will be enough; you should also inject in the abdomen a little amount of NBF also). Try to leave as much material in the front so you will get to cut to the lungs when trimming down the block. I recommend you also to process it enclosed in a cassette that keeps the structures in place, and to extend the times, not so much in the dehydrating as in the infiltrating steps. Had you been using the dehydrating/infiltrating steps I used to use (ethyl alcohol/isopropyl alcohol/mineral oil) you would get a better infiltration. Hope this will help. Rene J. Patsy Ruegg wrote: I am processing whole fetal mouse samples (these are pretty big, they have skin developed) and the investigator wants to have sections so that both lungs show up at the same time in the same section. Can someone recommend to me a way to dissect these to open them up so that they will fix and process well but still maintain the whole body architecture for sectioning. Thank you, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From naje1972 <@t> yahoo.com Wed Oct 5 16:14:29 2005 From: naje1972 <@t> yahoo.com (cynthia haynes) Date: Wed Oct 5 16:14:51 2005 Subject: [Histonet] 20 micron sections Message-ID: <20051005211429.83499.qmail@web33002.mail.mud.yahoo.com> Hello, everyone I am in need of information. I am cutting brain(human)tissue at 6 microns and 20 microns. The 6 microns sections are perfect. The 20 microns sections are an nightmare(and I mean Elm street nightmare). Can anyone suggest the best way to cut these sections without getting wrinkles and folds. I am at my wits end. Thanks in advance Cynthia Haynes H.T. From hhawkins <@t> UTMB.EDU Wed Oct 5 16:15:48 2005 From: hhawkins <@t> UTMB.EDU (Hawkins, Hal K.) Date: Wed Oct 5 16:16:02 2005 Subject: [Histonet] lymphatics Message-ID: <8D6F233E2A5D574B929F3944F3316FD006310469@EXCH2K3.utmb.edu> For a model of necrotizing enterocolitis in the mouse we need to distinguish small veins from lymphatics in the gut. Could someone remind me of any immunostains or lectin that will allow us to tell the two apart? Many thanks, Hal Hawkins, UTMB Galveston From ploykasek <@t> phenopath.com Wed Oct 5 16:53:42 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Wed Oct 5 16:54:19 2005 Subject: [Histonet] Pax2 Message-ID: Hi all. I have a question about an antibody to Pax2. We have just begun to try to work up this antibody. It is supposedly upregulated in renal cell carcinoma, so it should be positive & it has been. From the reading I have done it should be negative in normal adult kidney. Our current problem is that we are seeing staining on normal adult kidney. Any help or ideas are appreciated. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information.? Any unauthorized review, use, disclosure or distribution is prohibited.? If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message or you may call PhenoPath Laboratories, Seattle, WA U.S.A at (206) 374-9000. From dford <@t> deltapathology.com Wed Oct 5 17:21:44 2005 From: dford <@t> deltapathology.com (D. Ford) Date: Wed Oct 5 17:15:32 2005 Subject: [Histonet] RE: Positions in Louisiana In-Reply-To: Message-ID: Delta Pathology, in Shreveport Louisiana, currently looking for HT/HTLs. If you have been displaced by the hurricanes and want to return to your home state or if you are looking to relocate please contact us for more information. Darlene Ford, B.S. CT, HT (ASCP) Technical Supervisor 2915 Missouri Avenue Shreveport, LA 71109 318-621-8820 phone 318-671-5922 fax dford@deltapathology.com From LuckG <@t> empirehealth.org Wed Oct 5 17:44:14 2005 From: LuckG <@t> empirehealth.org (Luck, Greg D.) Date: Wed Oct 5 17:44:29 2005 Subject: [Histonet] lymphatics Message-ID: Hal, Why don't you try the tools offered on www.immunoquery.com (this one is the best but you need to get a username and password) or www.IHCWorld.com. These are both excellent IHC resources. Greg Luck, BS, HT(ASCP) Anatomic Pathology Supervisor Deaconess Medical Center 800 W. 5th Ave Spokane, WA 99204 Phone 509.473.7077 Fax 509.473.7133 luckg@empirehealth.org www.deaconessmedicalcenter.org -----Original Message----- From: Hawkins, Hal K. [mailto:hhawkins@UTMB.EDU] Sent: Wednesday, October 05, 2005 2:16 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] lymphatics For a model of necrotizing enterocolitis in the mouse we need to distinguish small veins from lymphatics in the gut. Could someone remind me of any immunostains or lectin that will allow us to tell the two apart? Many thanks, Hal Hawkins, UTMB Galveston _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From conniegrubaugh <@t> hotmail.com Wed Oct 5 22:57:47 2005 From: conniegrubaugh <@t> hotmail.com (connie grubaugh) Date: Wed Oct 5 22:58:11 2005 Subject: [Histonet] Nevada Society of Histotechnology Message-ID: The Nevada Society of HIstotechnology will be having their next seminar Oct. 28th and 29th at Sunrise Hospital Auditorium in Las Vegas Nevada. There will be a Friday evening get together with the vendors. Drinks and snacks will be provided. This will start at 5:30 pm. Saturday sign in at 7 am. Coffee and rolls will be provided by Fisher Scientific. Meeting starts at 8 am. First meeting will be Pam Marcum -Is Your Tissue Processor Fighting You? Fight Back. This will be 3 ceu"s. There will be a luncheon provided by Sakura and Cardinal Health. A business meeting will also be held at this time. Afternoon meetings. 1 pm- Sandra L. Cummings, Putting the bug in Histotechnology, A basic understanding of Microorganisms. To follow Marianne Mailhoit- Exposure control plan for Blood Borne Pathogens. There will be no charge for this meeting. Hope to see everyone there. Vegas is the best place to be on Halloween weekend. Please email me with any questions, or call 702-396-4079 or 702-938-3619. Connie Grubaugh Nevada Society of Histotechnology Striving for Excellence Despite the Odds. From jkiernan <@t> uwo.ca Wed Oct 5 23:18:11 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Wed Oct 5 23:18:37 2005 Subject: [Histonet] lymphatics References: <8D6F233E2A5D574B929F3944F3316FD006310469@EXCH2K3.utmb.edu> Message-ID: <4344A583.842FBFA@uwo.ca> Dear Hal Hawkins, The endothelium of lymphatic vessels can be stained by enzyme activity histochemistry. The enzyme is a 5-nucleotidase (belongs to the phosphatase family). Ordinarily this type of method is carried out with frozen sections of tissue that has been minimally fixed in formaldehyde or glutaraldehyde, or with unfixed cryostat sections that have been briefly fixed in cold acetone. The enzyme in lymphatics may be fairly robust because its activity can survive embedding in glycol methacrylate. The references below may help. If you choose one of these methods it will be important to consult the original publication because there's more than one sort of 5-nucleotidase. Each ref is followed by brief notes that I took when reading the papers. [An R in the acquisition number means I've got a Reprint of the whole paper; an A usually means I have a copy of the paper's Abstract. No letter with the acquisition number usually means I've seen the whole paper and took notes in the library.] 8194R. Kato,S; Yasunaga,A; Uchida,U (1991): Enzyme-histochemical method for identification of lymphatic capillaries. Lymphology 24, 125-129. Glycol methacrylate-embedded sections stained for 5'-nucleotidase & alkaline phosphatase. 5-N-ase in lymph capills; AlkP-ase in blood capills. 9239R. Nishida,S; Ohkuma,M (1993): Enzyme-histochemical staining of dermal lymphatic capillaries by guanylate cyclase. Lymphology 26, 195-199. Enzyme histochemical staining distinguishes lymph from blood capillaries. Says 5-nucleotidase & adenylate cyclase do so too. (Used unfixed cryostat sections.) 9665R. Okada,E (1994): An improved enzyme-histochemical method for identification of lymphatic capillaries on paraffin sections. Lymphology 27, Suppl, 732-735. Staining of blood & lymph capillaries. Enzyme histochemistry for 5-nucleotidase in presence of levamisole for lymph capills; alkaline phosphatase for blood capills. 11114A. Miura,M; Kato,S; von Ludinghausen,M (1998): Lymphatic drainage of the cerebrospinal fluid from monkey spinal meninges with special reference to the distribution of the epidural lymphatics. Arch. Histol. Cytol. 61(3, Aug), 277-286. 5'-nucleotidase staining for lymphatics; alk phosphatase for blood capillaries (Kato et al '91,'93). Also traced carbon particles from cisterna magna. Lymphatics and carbon found on surfaces of cervical & thoracic (most at brachial plexus levels) roots; not lumbosacral. Epidural lymphatics most developed on dorsal surface of lower cervical dura. I hope this helps. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Hawkins, Hal K." wrote: > > > For a model of necrotizing enterocolitis in the mouse we need to > distinguish small veins from lymphatics in the gut. Could someone > remind me of any immunostains or lectin that will allow us to tell the > two apart? > > Many thanks, > > Hal Hawkins, UTMB Galveston > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From anh2006 <@t> med.cornell.edu Wed Oct 5 23:57:15 2005 From: anh2006 <@t> med.cornell.edu (anh2006@med.cornell.edu) Date: Wed Oct 5 23:57:37 2005 Subject: [Histonet] lymphatics In-Reply-To: <4344A583.842FBFA@uwo.ca> References: <8D6F233E2A5D574B929F3944F3316FD006310469@EXCH2K3.utmb.edu> <4344A583.842FBFA@uwo.ca> Message-ID: <48830.66.108.123.192.1128574635.squirrel@webmail> I read Dr. Kiernan's post with much interest. Has anyone on Histonet had direct experience with these protocols? I would love to get some first hand experience feedback before I try it out. Thanks, Andrea > Dear Hal Hawkins, > > The endothelium of lymphatic vessels can be stained by > enzyme activity histochemistry. The enzyme is a > 5-nucleotidase (belongs to the phosphatase family). > Ordinarily this type of method is carried out with frozen > sections of tissue that has been minimally fixed in > formaldehyde or glutaraldehyde, or with unfixed cryostat > sections that have been briefly fixed in cold acetone. The > enzyme in lymphatics may be fairly robust because its > activity can survive embedding in glycol methacrylate. > > The references below may help. If you choose one of these > methods it will be important to consult the original > publication because there's more than one sort of > 5-nucleotidase. Each ref is followed by brief notes that I > took when reading the papers. [An R in the acquisition > number means I've got a Reprint of the whole paper; an A > usually means I have a copy of the paper's Abstract. No > letter with the acquisition number usually means I've seen > the whole paper and took notes in the library.] > > 8194R. Kato,S; Yasunaga,A; Uchida,U (1991): > Enzyme-histochemical method for identification of lymphatic > capillaries. Lymphology 24, 125-129. > Glycol methacrylate-embedded sections stained for > 5'-nucleotidase & alkaline phosphatase. 5-N-ase in lymph > capills; AlkP-ase in blood capills. > > 9239R. Nishida,S; Ohkuma,M (1993): Enzyme-histochemical > staining of dermal lymphatic capillaries by guanylate > cyclase. Lymphology 26, 195-199. > Enzyme histochemical staining distinguishes lymph from > blood capillaries. Says 5-nucleotidase & adenylate cyclase > do so too. (Used unfixed cryostat sections.) > > 9665R. Okada,E (1994): An improved enzyme-histochemical > method for identification of lymphatic capillaries on > paraffin sections. Lymphology 27, Suppl, 732-735. > Staining of blood & lymph capillaries. Enzyme > histochemistry for 5-nucleotidase in presence of levamisole > for lymph capills; alkaline phosphatase for blood capills. > > 11114A. Miura,M; Kato,S; von Ludinghausen,M (1998): > Lymphatic drainage of the cerebrospinal fluid from monkey > spinal meninges with special reference to the distribution > of the epidural lymphatics. Arch. Histol. Cytol. 61(3, Aug), > 277-286. > 5'-nucleotidase staining for lymphatics; alk phosphatase > for blood capillaries (Kato et al '91,'93). Also traced > carbon particles from cisterna magna. Lymphatics and carbon > found on surfaces of cervical & thoracic (most at brachial > plexus levels) roots; not lumbosacral. Epidural lymphatics > most developed on dorsal surface of lower cervical dura. > > I hope this helps. > -- > ------------------------------- > John A. Kiernan > Department of Anatomy and Cell Biology > The University of Western Ontario > London, Canada N6A 5C1 > kiernan[AT]uwo.ca > http://publish.uwo.ca/~jkiernan/ > http://instruct.uwo.ca/anatomy/530/index.htm From Tanni.Ahmed <@t> intervet.com Thu Oct 6 03:05:33 2005 From: Tanni.Ahmed <@t> intervet.com (Ahmed, T (Tanni)) Date: Thu Oct 6 03:09:28 2005 Subject: [Histonet] Fixation using Modified Bouins and NBF Message-ID: <09E7875E806DFB4BBECBADBF966BDBB6F6742B@mksn79.d50.intra> Dear Histonetters, I would like to know if fixation of avian bursa tissue with Modified Bouins fixative over 10% Neutral Buffered Formalin has any advantagous effect on immuno precipitation? Thanks in advance, Tanni Tanni S Ahmed Scientific Officer - Histopathology, R&D Intervet UK Ltd. Walton Manor, Walton, Milton Keynes, Buckinghamshire, MK7 7AJ, UK. Tel. +44(0)1908 685552/685543 Fax +44(0)1908 685614 E-mail: tanni.ahmed@intervet.com -------------------------------------- This message, including attachments, is confidential and may be privileged. If you are not an intended recipient, please notify the sender then delete and destroy the original message and all copies. You should not copy, forward and/or disclose this message, in whole or in part, without permission of the sender. -------------------------------------- From lpwenk <@t> sbcglobal.net Thu Oct 6 04:03:32 2005 From: lpwenk <@t> sbcglobal.net (Lee & Peggy Wenk) Date: Thu Oct 6 04:04:12 2005 Subject: [Histonet] 20 micron sections In-Reply-To: <20051005211429.83499.qmail@web33002.mail.mud.yahoo.com> Message-ID: Several suggestions: - Cut slower (real slow; real, real slow) - Cut the blocks at room temperature (not iced/chilled). Chilled is great for getting thiner sections, warmer for thicker sections - Might have to change the knife angle. Try cutting an "empty" paraffin block (make a block with no tissue), and increase and decrease the knife angle, until you can get sections without wrinkles from the plain block of paraffin. - If all the above fail, float the section in a container of 25% alcohol, then pick the section up on a slide, drain the slide of the alcohol, and slower lower the slide almost horizontal into the warm water flotation bath. The folds need to be parallel to the surface of the water bath, not perpendicular, in order for them to be pulled out by the difference in surface tension between the water and the alcohol. If 25% alcohol is ripping the section apart when laid on the water, try a lower percent of alcohol. Let us know what worked for you, so we all can learn. Peggy A. Wenk, HTL(ASCP)SLS William Beaumont Hospital Royal Oak, MI 48073 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of cynthia haynes Sent: Wednesday, October 05, 2005 5:14 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] 20 micron sections Hello, everyone I am in need of information. I am cutting brain(human)tissue at 6 microns and 20 microns. The 6 microns sections are perfect. The 20 microns sections are an nightmare(and I mean Elm street nightmare). Can anyone suggest the best way to cut these sections without getting wrinkles and folds. I am at my wits end. Thanks in advance Cynthia Haynes H.T. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Stephen.Eyres <@t> sanofi-aventis.com Thu Oct 6 04:26:11 2005 From: Stephen.Eyres <@t> sanofi-aventis.com (Stephen.Eyres@sanofi-aventis.com) Date: Thu Oct 6 04:26:27 2005 Subject: [Histonet] GFAP protocol for fluorescence Message-ID: Hi, Does anyone have a GFAP protocol for a fluorescence method that they could share? Many thanks Steve ---------------------------------------------------------- Le pr?sent message ainsi que ses ?ventuelles pi?ces jointes est exclusivement destin? au(x) destinataire(s), personnes physiques ou morales, qu'il d?signe. Il constitue de ce fait une correspondance ? caract?re priv? et peut contenir des informations confidentielles. Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser imm?diatement l'exp?diteur par retour de courrier ?lectronique puis de le d?truire, ainsi que ses ?ventuelles pi?ces jointes, sans en conserver de copie. This message, including any attachment, is intended for the use of the individual or entity to which it is addressed. It is therefore to be considered as a private correspondence which may contain confidential information. If you are not the intended recipient, please advise the sender immediately by reply e.mail and delete this message and any attachment thereto without retaining a copy. ---------------------------------------------------------- From BSauer <@t> stjosephswb.com Thu Oct 6 04:55:39 2005 From: BSauer <@t> stjosephswb.com (Sauer, Barb) Date: Thu Oct 6 04:55:57 2005 Subject: [Histonet] staining under hoods Message-ID: Does anyone use a Labconco (or another)hood to do staining of H&E's? We recently moved into a new facility and when we moved none of our stains worked right. We have not completely ruled out the water change but we now have purchased stain(instead of making our own) and the stain is good. It would really be helpful to get feedback on who has the entire staining set up under a hood in comparison to only the xylene under the hood. We also have had the problem of a Labconco hood that we purchased that is supposed to move the height to adjust to different size people using it. It's a great concept but Labconco did not supply the ventilation tubing to make it moveable, thus we are stuck with an expensive hood that doesn't move. Any ideas???? Thanks for your help. B. Sauer Histology Department Synergy Health / St. Joesph's Hospital West Bend, WI 53095 ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you have received this email in error please notify the system manager. This footnote also confirms that this email message has been swept by MIMEsweeper for the presence of computer viruses. www.mimesweeper.com ********************************************************************** From JWEEMS <@t> sjha.org Thu Oct 6 06:51:04 2005 From: JWEEMS <@t> sjha.org (Weems, Joyce) Date: Thu Oct 6 06:51:13 2005 Subject: [Histonet] staining under hoods Message-ID: <83AACDB0810528418AA106F9AE9B7F7E01304DE2@sjhaexc02.sjha.org> Can you change the vent tubing to flexible dryer vent tubing? -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Sauer, Barb Sent: Thursday, October 06, 2005 5:56 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] staining under hoods Does anyone use a Labconco (or another)hood to do staining of H&E's? We recently moved into a new facility and when we moved none of our stains worked right. We have not completely ruled out the water change but we now have purchased stain(instead of making our own) and the stain is good. It would really be helpful to get feedback on who has the entire staining set up under a hood in comparison to only the xylene under the hood. We also have had the problem of a Labconco hood that we purchased that is supposed to move the height to adjust to different size people using it. It's a great concept but Labconco did not supply the ventilation tubing to make it moveable, thus we are stuck with an expensive hood that doesn't move. Any ideas???? Thanks for your help. B. Sauer Histology Department Synergy Health / St. Joesph's Hospital West Bend, WI 53095 ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you have received this email in error please notify the system manager. This footnote also confirms that this email message has been swept by MIMEsweeper for the presence of computer viruses. www.mimesweeper.com ********************************************************************** _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet Confidentiality Notice ** The information contained in this message may be privileged and is confidential information intended for the use of the addressee listed above. If you are neither the intended recipient nor the employee or agent responsible for delivering this message to the intended recipient, you are hereby notified that any disclosure, copying, distribution or the taking of any action in reliance on the contents of this information is strictly prohibited. If you have received this communication in error, please notify us immediately by replying to the message and deleting it from your computer. Thank you. Saint Josephs Health System, Inc. From Ronnie_Houston <@t> bshsi.com Thu Oct 6 07:13:47 2005 From: Ronnie_Houston <@t> bshsi.com (Houston, Ronnie) Date: Thu Oct 6 07:14:10 2005 Subject: [Histonet] amyloid in FNA smears Message-ID: <530361BF03351B4CAE5270A05D3037B506B71A6E@bsrexms01.bshsir.com> What is the best fixation for smears from a fat-pad of a patient suspected with having amyloidosis, to get optimal preservation for demonstration of amyloid? Thanks Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com ________________________________________________________________________________________________________________________________ ________________________________________________________________________________________________________________________________ The information in this communication is intended to be confidential to the Individual(s) and/or Entity to whom it is addressed. It may contain information of a Privileged and/or Confidential nature, which is subject to Federal and/or State privacy regulations. In the event that you are not the intended recipient or the agent of the intended recipient, do not copy or use the information contained within this communication, or allow it to be read, copied or utilized in any manner, by any other person(s). Should this communication be received in error, please notify the sender immediately either by response e-mail or by phone, and permanently delete the original e-mail, attachment(s), and any copies. From danaspears <@t> frontiernet.net Thu Oct 6 07:14:55 2005 From: danaspears <@t> frontiernet.net (danaspears@frontiernet.net) Date: Thu Oct 6 07:15:14 2005 Subject: [Histonet] High Complexity Testing - IHC Message-ID: <20051006071455.wmqs0oc8scko4gw4@webmail.frontiernet.net> Hello everyone! I have been looking in the archives and am still a bit confused. I have a friend who has asked me to find out who may perform IHC in her lab... Would you say the following is correct as to who can perform IHC? BS or MT/HTL AS in science HT or MLT Prior to 24 April 1995 HS graduate AND Military training (50 wks) and classified as a Medical Laboratory Specialist OR Graduate of an accredited lab/histology program. Based on the above, I am interpreting that I currently have several techs that cannot perform IHC. Two are HT(ASCP), one HS/OJT the other GED/OJT. Neither with any college, military training, nor attended an accredited program. These two cannot perform the testing. I have two trainees, one has an associates, is an MLT, and is currently sitting for his HT. (I think he is okay, or does he have to wait to pass his HT?). The last is another trainee that has not completed her degree yet, so she has a ways to go yet. She will have her BS in May, then will sit for her HTL. Once she has her BS, she meets the requirements. Does all of this sound correct? Thanks for any and all help! From gillian.2.brown <@t> gsk.com Thu Oct 6 07:53:48 2005 From: gillian.2.brown <@t> gsk.com (gillian.2.brown@gsk.com) Date: Thu Oct 6 07:53:48 2005 Subject: [Histonet] Metachromasia of mast cells in epoxy resin sections Message-ID: Hi Histonetters, would any one have experience of counting granules in mouse mast cells in tissues which have been formaldehyde fixed, decalcified in a formic acid based solution, post osmicated and embedded in epoxy resin? I'm taking 1 micron sections and looking to assess the 'degranulation' status of each mast cell in terms of how many are still dark blue and how many are pink ie based on the metachromatic properties of toluidine blue. I suppose my question is will the formic acid have altered the dye binding properties so that I'll get erroneous results? I'm going to experiment with tissue that does not normally have this step but any insights now would be most welcome. Many thanks Gill Brown GlaxoSmithKline Medicines Research Centre, STEVENAGE, From Andrew.Prior <@t> Smith-Nephew.com Thu Oct 6 07:57:11 2005 From: Andrew.Prior <@t> Smith-Nephew.com (Prior, Andrew) Date: Thu Oct 6 07:58:33 2005 Subject: [Histonet] Histology Journals Message-ID: I was wondering if anyone could recommend a good histology journal, that covers a lot of the methods and equipment. I've had a look at some journals on the web, but would appreciate some feedback from regular readers on how useful you find them. Many thanks Andrew Andrew Prior Histologist Smith & Nephew Research Centre York Science Park Heslington York, YO10 5DF UK Andrew.Prior@smith-nephew.com Confidentiality. This electronic transmission is strictly confidential to Smith & Nephew and intended solely for the addressee. It may contain information which is covered by legal, professional or other privilege. If you are not the intended addressee, or someone authorised by the intended addressee to receive transmissions on behalf of the addressee, you must not retain, disclose in any form, copy or take any action in reliance on this transmission. If you have received this transmission in error, please notify the sender as soon as possible and destroy this message. From rjbuesa <@t> yahoo.com Thu Oct 6 08:25:24 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 6 08:25:44 2005 Subject: [Histonet] 20 micron sections In-Reply-To: <20051005211429.83499.qmail@web33002.mail.mud.yahoo.com> Message-ID: <20051006132524.26581.qmail@web61220.mail.yahoo.com> Try increasing the cutting angle; cutting more slow and cooling the cutting surface inmediately before the section you are going to take. cynthia haynes wrote:Hello, everyone I am in need of information. I am cutting brain(human)tissue at 6 microns and 20 microns. The 6 microns sections are perfect. The 20 microns sections are an nightmare(and I mean Elm street nightmare). Can anyone suggest the best way to cut these sections without getting wrinkles and folds. I am at my wits end. Thanks in advance Cynthia Haynes H.T. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From rjbuesa <@t> yahoo.com Thu Oct 6 08:32:56 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 6 08:33:09 2005 Subject: [Histonet] Histology Journals In-Reply-To: Message-ID: <20051006133256.73707.qmail@web61219.mail.yahoo.com> Although it could be considered as a "biassed" opinion I would suggest for you to look at the Journal of Histotechnology from the National Society of Histotechnology. Rene J. "Prior, Andrew" wrote: I was wondering if anyone could recommend a good histology journal, that covers a lot of the methods and equipment. I've had a look at some journals on the web, but would appreciate some feedback from regular readers on how useful you find them. Many thanks Andrew Andrew Prior Histologist Smith & Nephew Research Centre York Science Park Heslington York, YO10 5DF UK Andrew.Prior@smith-nephew.com Confidentiality. This electronic transmission is strictly confidential to Smith & Nephew and intended solely for the addressee. It may contain information which is covered by legal, professional or other privilege. If you are not the intended addressee, or someone authorised by the intended addressee to receive transmissions on behalf of the addressee, you must not retain, disclose in any form, copy or take any action in reliance on this transmission. If you have received this transmission in error, please notify the sender as soon as possible and destroy this message. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From rjbuesa <@t> yahoo.com Thu Oct 6 08:41:49 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 6 08:42:00 2005 Subject: [Histonet] Nevada Society of Histotechnology In-Reply-To: Message-ID: <20051006134149.25590.qmail@web61225.mail.yahoo.com> Connie: Could you use this oportunity to ask the members of the Nevada Society to contribute with Tricks of the Trade to be shared by all the fellow histotechs? This could be a great help for all! Rene J. Buesa connie grubaugh wrote: The Nevada Society of HIstotechnology will be having their next seminar Oct. 28th and 29th at Sunrise Hospital Auditorium in Las Vegas Nevada. There will be a Friday evening get together with the vendors. Drinks and snacks will be provided. This will start at 5:30 pm. Saturday sign in at 7 am. Coffee and rolls will be provided by Fisher Scientific. Meeting starts at 8 am. First meeting will be Pam Marcum -Is Your Tissue Processor Fighting You? Fight Back. This will be 3 ceu"s. There will be a luncheon provided by Sakura and Cardinal Health. A business meeting will also be held at this time. Afternoon meetings. 1 pm- Sandra L. Cummings, Putting the bug in Histotechnology, A basic understanding of Microorganisms. To follow Marianne Mailhoit- Exposure control plan for Blood Borne Pathogens. There will be no charge for this meeting. Hope to see everyone there. Vegas is the best place to be on Halloween weekend. Please email me with any questions, or call 702-396-4079 or 702-938-3619. Connie Grubaugh Nevada Society of Histotechnology Striving for Excellence Despite the Odds. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From Debbiejsiena <@t> aol.com Thu Oct 6 08:55:39 2005 From: Debbiejsiena <@t> aol.com (Debbiejsiena@aol.com) Date: Thu Oct 6 08:57:38 2005 Subject: [Histonet] Job Openings In Dallas, Texas Message-ID: Good Morning All Tissue Techniques In Dallas, Texas has several HT openings along with an opening for a Lab tech. If you are the best and expect the best of Histology personnel than this is the lab for you. We have work that allows people to be creative and solve problems along with doing some rearch projects.If you would like to be part of a winning team, please contact us. Debbie Siena, cell: 520-360-3013, Fred Siena, cell: 817-228-8023, Office: 972-241-6277, Fax 972-241-4747, e-mail: tissuetech@juno.com From Sue.Kapoor <@t> uhsi.org Thu Oct 6 08:56:14 2005 From: Sue.Kapoor <@t> uhsi.org (Kapoor, Sue) Date: Thu Oct 6 08:59:32 2005 Subject: [Histonet] CAP question Message-ID: <61E9F2400F53D5119CFC00508B44E33B02E9E1B7@khmcexch.uhsi.org> Hi everyone, is anyone familiar with CAP #ANP.22925 "For IHC tests that provide independent predictive/prognostic information, does the patient report include info on specimen processing, the antibody clone, and the scoring method used? (e.g., hormone receptor in breast carcinoma, HER-2/neu, EGFR). "The lab should periodically compare its patient results with published benchmarks, and also evaluate interobserver variabitlity among the pathologists in the lab". how are others handling this?? Many, many thanks in advance, Sue Kapoor, HT (ASCP) Histology Coordinator Kenosha Medical Center Kenosha, WI 262-653-5570 From pmarcum <@t> vet.upenn.edu Thu Oct 6 09:22:25 2005 From: pmarcum <@t> vet.upenn.edu (Pamela Marcum) Date: Thu Oct 6 09:22:55 2005 Subject: [Histonet] Region II Meeting November 4th and 5th, 2005 Message-ID: <6.1.1.1.2.20051006101724.01950860@mail.vet.upenn.edu> Region II is having a meeting!! We are including a copy of the program for your review. If anyone would like a copy of the full program with registration, cost, and directions please contact me. The meeting is at the Hilton Valley Forge Hotel in King of Prussia, PA - outside Philadelphia, PA. Best Regards, Pamela A Marcum Manager, Histology Special Procedures University of Pennsylvania School of Veterinary Medicine R.S. Reynolds Jr. CORL New Bolton Center 382 West Street Road Kennett Square, PA 19348 Phone - 610-925-6278 Fax - 610-925-8120 E-mail - pmarcum@vet.upenn.edu Friday - November 4, 2005 ? Region II NSH Fall Meeting Registration 7:00AM to 8:00AM WS #1- 8:00 to 11:30AM Muscle talk! Barone WS #2- 8:00 to 11:30AM Preparing for the IHC Qualification Examination (ASCP Registry) Macrea WS #3- 8:00 to 11:30AM Unlocking The Secrets of Mohs? Grossing and Cryosectioning Praet WS #4- 8:00 to 9:30AM Chemicon?s Advanced Tissue Arrayer (ATA 100), tissue Microarrays, IHC Select Reagents, and IHC Manual Select Staining for High Through-Put Screening Studies Schaub Morning Break In Exhibits ? 9:30 to 10:00AM WS # 5- 10:00 to 11:30AM Confocal, Multi-Photon and Deconvolution Microscopy in Clinical Diagnosis Atkins WS #6- 10:00 to 11:30AM Microwave Tissue Fixation And Processing Frei Afternoon Workshops and Seminars WS #7- 1:00 to 2:30PM Laboratory Management: Problems and Solutions Billings WS#8- 1:00 to 2:30PM What Every Histotech Should Know About Water Macrea Afternoon Break In Exhibits ? 2:30 to 3:00PM WS#9- 1:00 to 4:30PM Quality Assessment of Special Stains Micciche WS#10- 1:00 to 4:30PM From Amygdala to Arabidopsis: Why?s and How?s of Vibrating Blade Microscopy Freeland WS#11- 3:00 to 4:30PM Mummies: Ancient to Modern Wade WS#12- 3:00 to 4:30PM Reagent Alcohol ? Can?t Drink It ? So What Is It? Marcum Saturday - November 5, 2005 ? Region II NSH Fall Meeting Registration 7:00AM to 8:00AM WS#13- 8:00 to 9:30AM Are All Tissues Created Equal? Hughes Research vs Clinical Louro WS#14- 8:00 to 9:30AM High Profile Cases Let the Stress Begin Wetli WS#15- 8:00 to 11:30AM Identification of Multiple Antigens in the Same Specimen with Overview of Heat Induced Epitope Retrieval Procedure (HIER) Myers WS#16- 8:00AM to 4:30PM HT(ASCP) Readiness Examination Micciche Morning Break In Exhibits ? 9:30 to 10:00AM WS#17- 10:00 to 11:30AM Flow Cytometry Versus Immunohistochemistry ? When Is One Better? Wilson WS#18- 10:00 to 11:30AM Forensic Entomology D?Amico & D?Amico Afternoon Workshops and Seminars WS#19- 1:00 to 2:30PM History of Histology Mitchell WS#20- 1:00 to 4:30PM Reality of Forensic Science Desiderio New Jersey State Police Crime Laboratory Matukonis & Singh WS#21- 1:00 to 2:30PM What Is Your Body Saying To Me? Billings Afternoon Break NO Exhibits ? 2:30 to 3:00PM WS#22- 3:00 to 4:30PM Utilizing Performance Appraisal As A Motivating Tool Horstmann WS#23- 3:00 to 4:30PM Introduction to Fine Needle Aspiration for the Histotechnician Braud From gu.lang <@t> gmx.at Thu Oct 6 09:46:12 2005 From: gu.lang <@t> gmx.at (Gudrun Lang) Date: Thu Oct 6 09:46:35 2005 Subject: AW: [Histonet] lymphatics In-Reply-To: <8D6F233E2A5D574B929F3944F3316FD006310469@EXCH2K3.utmb.edu> Message-ID: I think, Podoplanin is typical for lymphatic endothelcells. We purchase ours from Acris, Germany. Gudrun Lang -----Urspr?ngliche Nachricht----- Von: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] Im Auftrag von Hawkins, Hal K. Gesendet: Mittwoch, 05. Oktober 2005 23:16 An: Histonet@lists.utsouthwestern.edu Betreff: [Histonet] lymphatics For a model of necrotizing enterocolitis in the mouse we need to distinguish small veins from lymphatics in the gut. Could someone remind me of any immunostains or lectin that will allow us to tell the two apart? Many thanks, Hal Hawkins, UTMB Galveston _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rjbuesa <@t> yahoo.com Thu Oct 6 09:57:03 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 6 09:57:20 2005 Subject: [Histonet] CAP question In-Reply-To: <61E9F2400F53D5119CFC00508B44E33B02E9E1B7@khmcexch.uhsi.org> Message-ID: <20051006145703.41370.qmail@web61225.mail.yahoo.com> In our lab we used to mention the antibody, the scoring method, but not the procedure.Any additional information depends on the pathologist's desire about being specific on the procedure. We never had any problems during a CAP inspection. Rene J. "Kapoor, Sue" wrote: Hi everyone, is anyone familiar with CAP #ANP.22925 "For IHC tests that provide independent predictive/prognostic information, does the patient report include info on specimen processing, the antibody clone, and the scoring method used? (e.g., hormone receptor in breast carcinoma, HER-2/neu, EGFR). "The lab should periodically compare its patient results with published benchmarks, and also evaluate interobserver variabitlity among the pathologists in the lab". how are others handling this?? Many, many thanks in advance, Sue Kapoor, HT (ASCP) Histology Coordinator Kenosha Medical Center Kenosha, WI 262-653-5570 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From anh2006 <@t> med.cornell.edu Thu Oct 6 10:10:55 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Thu Oct 6 10:06:32 2005 Subject: [Histonet] Cryostats Message-ID: Hello everyone ... What is everyone's favorite cryostat and why? We are in the market. I have a few personal favorites but I wanted to hear from the general audience what is liked best. We cut mostly mouse tissue (probably 70-80% mouse tissue) and 50% is probably mouse bone. Thanks! Andrea -- From anh2006 <@t> med.cornell.edu Thu Oct 6 10:11:55 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Thu Oct 6 10:07:34 2005 Subject: AW: [Histonet] lymphatics In-Reply-To: <200510061447.j96ElFoh222410@smtp-in1.med.cornell.edu> References: <200510061447.j96ElFoh222410@smtp-in1.med.cornell.edu> Message-ID: In the mouse I do not find podoplanin to be specific for lymphatics. I gave up on it as it also clearly stained some other fibres in the tumor sections we were examining. This could be antibody dependent as I only tried it from one source, although a well established source. I know believe that Lyve-1+/Prox-1+ doublestaining is the way to go. I believe lymphatics are currenlty being described as Lyve-1+/Prox-1+/VEGFR-3+/Podoplanin+ although none of these markers can be used exclusively for identification - at least in the mouse. At 4:46 PM +0200 10/6/05, Gudrun Lang wrote: >I think, Podoplanin is typical for lymphatic endothelcells. We purchase ours >from Acris, Germany. > >Gudrun Lang -- From BDUE <@t> PARTNERS.ORG Thu Oct 6 10:09:42 2005 From: BDUE <@t> PARTNERS.ORG (Due, Brice) Date: Thu Oct 6 10:09:53 2005 Subject: [Histonet] 20 micron sections Message-ID: <59C772E2D8EDF345AE1601F5B60B3CB501027194@PHSXMB7.partners.org> Hello Cynthia, we cut thick (10-20um) paraffin brain here fairly often. Someone else already mentioned that warmer is better. I find 10um cuts well at room temp., however 20um cuts even better if I throw the block into the hot water bath for a few seconds to soften it up. Or I soak a gauze in the hot water and press it to the block face (in the chuck) periodically. The temperature issue usually causes chatter/shatter artifacts. If you're getting wrinkles on thick sections, I'm guessing it's from the swell on the water bath. Your perfect flat 20um section becomes "rippled" when the grey & white matters swell differentially on the hot water. Then it won't lay flat on the slide. You can try increasing the temp of your water bath to spread the section more. Or you can lower the temp alot to reduce the swell. Neither will eliminate the problem all the time. Above 15um, it becomes more difficult to get good adhesion. Seemingly this is because the thick sections trap water more effectively and/or don't lay flat on the slide. Be sure you air dry the section *thoroughly* before melting them in a hot oven. We usually put sections in a 37C oven overnight and melt at 60C the next day. To deal with the wrinkle/adhesion problem, here's a gem from the old-timer who taught me. Place a few pieces of filter paper on a bench surface. Soak it with some 95-100% ethanol. Not just damp, but soaking dripping wet. Cut your section and pick it up off a hot water bath. It will not lay flat on the slide, but strive for the flattest you can. Stand the slide up for 1-5min to drain some of the water. Here's the trick. Now carefully lay the slide **section DOWN** onto the wet filter paper. Hold the slide firmly in place and rub down the back side hard. I was taught to rub in only one direction (pick one), but I've found that rubbing randomly works just as well. Now *carefully* pick up the slide from the paper. I've found that the easiest way to do this is to pull the filter paper to the edge of the bench and pull the paper downwards. Magically the section stays on the slide. If it sticks to the paper, use more ethanol before laying the section down or use a clean paper on top. Change filter paper if paraffin fragments stick, or every several slides. This trick works well with 20um and thicker. For sections thinner than 15um I've never needed it. Note, this will force/produce some wrinkles because of the squishing effect, but most of the section will get flattened out onto the slide, with good adhesion. We do this for large sections of brain (up to 6"x8"). If you can use 15um sections, they are much easier to cut and work with. They flatten out better and have fewer drying issues. When I jump from 15um to 20um I usually have to dig into my bag of tricks. Best of Luck, -brice Neuropathology Lab Brigham & Women's Hospital Boston, MA "THE INFORMATION TRANSMITTED IN THIS EMAIL IS INTENDED ONLY FOR THE PERSON OR ENTITY TO WHICH IT IS ADDRESSED AND MAY CONTAIN CONFIDENTIAL AND/OR PRIVILEGED MATERIAL. ANY REVIEW, RETRANSMISSION, DISSEMINATION OR OTHER USE OF OR TAKING OF ANY ACTION IN RELIANCE UPON, THIS INFORMATION BY PERSONS OR ENTITIES OTHER THAN THE INTENDED RECIPIENT IS PROHIBITED. IF YOU RECEIVE THIS EMAIL IN ERROR, PLEASE CONTACT THE SENDER AND DELETE THE MATERIAL FROM ANY COMPUTER." HIPPA Policy, 2003, Brigham & Women's Hospital, Boston, MA -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of cynthia haynes Sent: Wednesday, October 05, 2005 5:14 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] 20 micron sections Hello, everyone I am in need of information. I am cutting brain(human)tissue at 6 microns and 20 microns. The 6 microns sections are perfect. The 20 microns sections are an nightmare(and I mean Elm street nightmare). Can anyone suggest the best way to cut these sections without getting wrinkles and folds. I am at my wits end. Thanks in advance Cynthia Haynes H.T. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From ian.montgomery <@t> bio.gla.ac.uk Thu Oct 6 10:20:50 2005 From: ian.montgomery <@t> bio.gla.ac.uk (Ian Montgomery) Date: Thu Oct 6 10:21:07 2005 Subject: Fwd: [Histonet] Cryostats Message-ID: <6.2.3.4.2.20051006161247.030f2b70@udcf.gla.ac.uk> Andrea, I bought my Bright cryostat in 1974 and in my opinion it's the Rolls Royce of cryostats. 31 years of constant use, never been out of service and cuts as good today as back in 74. Ian. >Hello everyone ... > >What is everyone's favorite cryostat and why? We are in the market. >I have a few personal favorites but I wanted to hear from the >general audience what is liked best. > >We cut mostly mouse tissue (probably 70-80% mouse tissue) and 50% is >probably mouse bone. > >Thanks! >Andrea >-- > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Dr. Ian Montgomery, Histotechnology, IBLS Support Services, Graham Kerr Building, Institute of Biomedical & Life Sciences, University of Glasgow, Glasgow, G12 8QQ. Tel: 0141 339 8855 Office: 4652 Lab: 6644. Pager: 07623 975451 e-mail: ian.montgomery@bio.gla.ac.uk From ian.montgomery <@t> bio.gla.ac.uk Thu Oct 6 10:35:57 2005 From: ian.montgomery <@t> bio.gla.ac.uk (Ian Montgomery) Date: Thu Oct 6 10:36:11 2005 Subject: Fwd: RE: [Histonet] Cryostats Message-ID: <6.2.3.4.2.20051006163137.030df730@udcf.gla.ac.uk> Fair's fair, praise where it's due. When I was a boy I was brought up on SLEE cryostats so Bright was a novelty at the time. Over the years I've used many cryostat but as with the ladies, I always return to my first love. Ian. >Alan must love you too:-) > >Dr Terry L Marshall, B.A.(Law), M.B.,Ch.B.,F.R.C.Path > Consultant Pathologist > Rotherham General Hospital > South Yorkshire > England > terry.marshall@rothgen.nhs.uk > >-----Original Message----- >From: Ian Montgomery [mailto:ian.montgomery@bio.gla.ac.uk] >Sent: 06 October 2005 16:21 >To: histonet@lists.utsouthwestern.edu >Subject: Fwd: [Histonet] Cryostats > > >Andrea, > I bought my Bright cryostat in 1974 and in my opinion it's >the Rolls Royce of cryostats. 31 years of constant use, never been >out of service and cuts as good today as back in 74. >Ian. > > > > >Hello everyone ... > > > >What is everyone's favorite cryostat and why? We are in the market. > >I have a few personal favorites but I wanted to hear from the > >general audience what is liked best. > > > >We cut mostly mouse tissue (probably 70-80% mouse tissue) and 50% is > >probably mouse bone. > > > >Thanks! > >Andrea > >-- > > > >_______________________________________________ > >Histonet mailing list > >Histonet@lists.utsouthwestern.edu > >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > >Dr. Ian Montgomery, >Histotechnology, >IBLS Support Services, >Graham Kerr Building, >Institute of Biomedical & Life Sciences, >University of Glasgow, >Glasgow, >G12 8QQ. >Tel: 0141 339 8855 >Office: 4652 >Lab: 6644. >Pager: 07623 975451 >e-mail: ian.montgomery@bio.gla.ac.uk > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Dr. Ian Montgomery, Histotechnology, IBLS Support Services, Graham Kerr Building, Institute of Biomedical & Life Sciences, University of Glasgow, Glasgow, G12 8QQ. Tel: 0141 339 8855 Office: 4652 Lab: 6644. Pager: 07623 975451 e-mail: ian.montgomery@bio.gla.ac.uk From kgibbon <@t> qltinc.com Thu Oct 6 10:47:05 2005 From: kgibbon <@t> qltinc.com (kgibbon@qltinc.com) Date: Thu Oct 6 10:47:19 2005 Subject: Fwd: RE: [Histonet] Cryostats Message-ID: I too was brought up on SLEE cryostats and thought they were the best, but over the pond they were difficult to get so I use a Microm these days and would not change now. That automatic feed is great and with that addition of a "cryojane" the toughest tissues are nice to cut. Kevin Gibbon QLT Inc. |---------+-------------------------------------------> | | "Ian Montgomery" | | | | | | Sent by: | | | | | | | | | | | | 10/06/2005 08:35 AM | | | | |---------+-------------------------------------------> >------------------------------------------------------------------------------------------------------------------------------| | | | To: | | cc: | | Subject: Fwd: RE: [Histonet] Cryostats | >------------------------------------------------------------------------------------------------------------------------------| Fair's fair, praise where it's due. When I was a boy I was brought up on SLEE cryostats so Bright was a novelty at the time. Over the years I've used many cryostat but as with the ladies, I always return to my first love. Ian. >Alan must love you too:-) > >Dr Terry L Marshall, B.A.(Law), M.B.,Ch.B.,F.R.C.Path > Consultant Pathologist > Rotherham General Hospital > South Yorkshire > England > terry.marshall@rothgen.nhs.uk > >-----Original Message----- >From: Ian Montgomery [mailto:ian.montgomery@bio.gla.ac.uk] >Sent: 06 October 2005 16:21 >To: histonet@lists.utsouthwestern.edu >Subject: Fwd: [Histonet] Cryostats > > >Andrea, > I bought my Bright cryostat in 1974 and in my opinion it's >the Rolls Royce of cryostats. 31 years of constant use, never been >out of service and cuts as good today as back in 74. >Ian. > > > > >Hello everyone ... > > > >What is everyone's favorite cryostat and why? We are in the market. > >I have a few personal favorites but I wanted to hear from the > >general audience what is liked best. > > > >We cut mostly mouse tissue (probably 70-80% mouse tissue) and 50% is > >probably mouse bone. > > > >Thanks! > >Andrea > >-- > > > >_______________________________________________ > >Histonet mailing list > >Histonet@lists.utsouthwestern.edu > >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > >Dr. Ian Montgomery, >Histotechnology, >IBLS Support Services, >Graham Kerr Building, >Institute of Biomedical & Life Sciences, >University of Glasgow, >Glasgow, >G12 8QQ. >Tel: 0141 339 8855 >Office: 4652 >Lab: 6644. >Pager: 07623 975451 >e-mail: ian.montgomery@bio.gla.ac.uk > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Dr. Ian Montgomery, Histotechnology, IBLS Support Services, Graham Kerr Building, Institute of Biomedical & Life Sciences, University of Glasgow, Glasgow, G12 8QQ. Tel: 0141 339 8855 Office: 4652 Lab: 6644. Pager: 07623 975451 e-mail: ian.montgomery@bio.gla.ac.uk _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This electronic transmission (including any and all attachments) is intended solely for the use of the individual or entity to whom it is addressed and may contain information that is privileged and/or confidential. If you are not the intended recipient of this electronic transmission, you are hereby notified that any disclosure, copying or distribution, or the taking of any action in reliance upon the contents of this electronic transmission, is strictly prohibited, and you are further requested to purge this electronic transmission and all copies thereof from your computer system. From jkiernan <@t> uwo.ca Thu Oct 6 10:51:55 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Thu Oct 6 10:52:14 2005 Subject: [Histonet] Fixation using Modified Bouins and NBF References: <09E7875E806DFB4BBECBADBF966BDBB6F6742B@mksn79.d50.intra> Message-ID: <4345481B.9DABCFC7@uwo.ca> What is the composition of the "modified Bouin's" fixative, and how long did the specimens remain in it? Back in the old days Bouin's was considered a good fixative for some immunofluorescent applications, especially localizing protein and peptide hormones. The picric acid in the solution subdues much of the autofluorescence. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Ahmed, T (Tanni)" wrote: > > > Dear Histonetters, > > I would like to know if fixation of avian bursa tissue with Modified > Bouins fixative over 10% Neutral Buffered Formalin has any advantagous > effect on immuno precipitation? > > Thanks in advance, > > Tanni > > > > > > Tanni S Ahmed > > Scientific Officer - Histopathology, R&D > > > > Intervet UK Ltd. > > Walton Manor, > > Walton, Milton Keynes, > > Buckinghamshire, > > MK7 7AJ, UK. > > > > Tel. +44(0)1908 685552/685543 > > Fax +44(0)1908 685614 > > E-mail: tanni.ahmed@intervet.com > > > > > > > > -------------------------------------- > This message, including attachments, is confidential and may be privileged. > If you are not an intended recipient, please notify the sender then delete > and destroy the original message and all copies. You should not copy, forward > and/or disclose this message, in whole or in part, without permission of > the sender. > -------------------------------------- > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From asmith <@t> mail.barry.edu Thu Oct 6 11:20:40 2005 From: asmith <@t> mail.barry.edu (Smith, Allen) Date: Thu Oct 6 11:21:18 2005 Subject: [Histonet] Histology Journals Message-ID: <5D2189E74151CC42BEC02906BA8996322B910A@exchsrv01.barrynet.barry.edu> Our field is full of small-circulation journals rather than having one dominant journal. I have found the following useful: Biotechnic and Histochemistry, Journal of Histotechnology, Histochemistry, American Journal of Clinical Pathology, Micron, Journal of Histochemistry and Cytochemistry. Allen A. Smith, Ph.D. Professor of Anatomy Barry University School of Graduate Medical Sciences Podiatric Medicine and Surgery Miami Shores, Florida 33161 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Prior, Andrew Sent: Thursday, October 06, 2005 8:57 AM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] Histology Journals I was wondering if anyone could recommend a good histology journal, that covers a lot of the methods and equipment. I've had a look at some journals on the web, but would appreciate some feedback from regular readers on how useful you find them. Many thanks Andrew Andrew Prior Histologist Smith & Nephew Research Centre York Science Park Heslington York, YO10 5DF UK Andrew.Prior@smith-nephew.com Confidentiality. This electronic transmission is strictly confidential to Smith & Nephew and intended solely for the addressee. It may contain information which is covered by legal, professional or other privilege. If you are not the intended addressee, or someone authorised by the intended addressee to receive transmissions on behalf of the addressee, you must not retain, disclose in any form, copy or take any action in reliance on this transmission. If you have received this transmission in error, please notify the sender as soon as possible and destroy this message. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet The information transmitted is intended only for the person or entity to which it is addressed and may contain confidential, and/or privileged material. No confidentiality or privilege is waived or lost by any errant transmission. If you receive this message in error, please immediately delete it and all copies of it from your system and notify the sender. E-mail transmission cannot be guaranteed to be secure or error-free as information could be intercepted, corrupted, lost, destroyed, arrive late or incomplete, or contain viruses. Barry University - Miami Shores, FL (http://www.barry.edu) From sharon.osborn <@t> dnax.org Thu Oct 6 11:23:36 2005 From: sharon.osborn <@t> dnax.org (Osborn, Sharon) Date: Thu Oct 6 11:24:13 2005 Subject: [Histonet] Journals Message-ID: <29B25753F6B1D51196110002A589D44402398214@PALMSG30.us.schp.com> Additional resources are the Histologics that Vinnie edits for Sakura (go to their website for archived and current copies); regional and state socitety newsletters which often have TOT's in them plus doing Google searches for various procedures which bring up some universities with websites aout histology. Some vendors also have good examples of stain technics (Sigma for one). I have a set of old texts which I have collected from clearing out closed down laboratories, offices, etc that are a great resource when I go to research some technics. I also enjoy knowing how a technic came about. You also have excellent sources in histonet contributors to point you to specific references and articles. sharon osborn DNAX, SP BioPharma Palo Alto, CA 650.496.6539 Message: 21 Date: Thu, 6 Oct 2005 06:32:56 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] Histology Journals To: "Prior, Andrew" , "'histonet@lists.utsouthwestern.edu'" Message-ID: <20051006133256.73707.qmail@web61219.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Although it could be considered as a "biassed" opinion I would suggest for you to look at the Journal of Histotechnology from the National Society of Histotechnology. Rene J. "Prior, Andrew" wrote: I was wondering if anyone could recommend a good histology journal, that covers a lot of the methods and equipment. I've had a look at some journals on the web, but would appreciate some feedback from regular readers on how useful you find them. Many thanks Andrew Andrew Prior Histologist Smith & Nephew Research Centre York Science Park Heslington York, YO10 5DF UK Andrew.Prior@smith-nephew.com ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. From jkiernan <@t> uwo.ca Thu Oct 6 12:23:40 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Thu Oct 6 12:23:57 2005 Subject: [Histonet] Histology Journals References: Message-ID: <43455D9C.41C83BE4@uwo.ca> The best value for money are the journals that come with society memberships (the first 3 in the following list). Journal of Histotechnology. 4 issues a year, with N.S.H. subscription ($40) http://www.nsh.org/membership/application.html Biotechnic & Histochemistry. 6 issues a year, with Biological Stain Commission subscription ($40) Membership is open to anyone with an interest in staining. Contact the Secretary, dpenney[AT]biostains.org (The application form at http://www.biostains.org/new_page_15.htm is out of date; you no longer need to be nominated by a member to join the BSC.) Journal of Histochemistry and Cytochemistry. 12 issues a year, with Histochemical Society subscription ($50) http://www.histochemicalsociety.org/members/application.shtml This has the highest impact factor in the field. Histochemistry and Cell Biology is another high impact (for the field) journal published by Springer for the Society for Histochemistry (a European society). I suspect it's too costly for most individuals to take, but it's in libraries. 12 issues a year. Journal of Molecular Histology (it was The Histochemical Journal until 2004) is now also published by Springer; 12 issues per year. Formerly was an option with subscription to the Royal Microscopical Society (?46). I don't know how much it costs now. Libraries have it. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Prior, Andrew" wrote: > > I was wondering if anyone could recommend a good histology journal, that > covers a lot of the methods and equipment. > I've had a look at some journals on the web, but would appreciate some > feedback from regular readers on how useful you find them. > > Many thanks > > Andrew > > Andrew Prior > Histologist > Smith & Nephew Research Centre > York Science Park > Heslington > York, YO10 5DF > UK > > Andrew.Prior@smith-nephew.com > > Confidentiality. > This electronic transmission is strictly confidential to Smith & Nephew and > intended solely for the addressee. It may contain information which is > covered by legal, professional or other privilege. If you are not the > intended addressee, or someone authorised by the intended addressee to > receive transmissions on behalf of the addressee, you must not retain, > disclose in any form, copy or take any action in reliance on this > transmission. If you have received this transmission in error, please notify > the sender as soon as possible and destroy this message. > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From liz <@t> premierlab.com Thu Oct 6 12:30:06 2005 From: liz <@t> premierlab.com (Elizabeth Chlipala) Date: Thu Oct 6 12:30:29 2005 Subject: [Histonet] labeling cells in culture Message-ID: <000201c5ca9b$981d0ef0$a7d48a80@AMY> Hello everyone I have a question that one of my clients has asked. Thanks in advance. Below is the question. Is there any method for labeling living cells in order to track them in mixtures with non-labeled cells? The point is that I would like to label primary human chondrocytes (difficult to transfect), culture them for 4 weeks together with non-labeled cells and then I would like to be able to look at these chondrocytes by microscopy. I know about various compounds (Quantum dots, etc) but those are good only for about one week. Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC P.O. Box 18592 Boulder, Colorado 80308 Office: (303) 735-5001 Fax: (303) 735-3540 liz@premierlab.com www.premierlab.com Ship to Address: Premier Laboratory University of Colorado MCDB, Room A3B40 Boulder, Colorado 80309 From donald <@t> labcareer.com Thu Oct 6 12:31:46 2005 From: donald <@t> labcareer.com (Donald Knowles) Date: Thu Oct 6 12:30:53 2005 Subject: [Histonet] Histotology Travel Position in Fl. Message-ID: <26A7728EF768DA478F45954252293C203A4C4C@matrix.HCCI.local> I have an urgent need for a person looking to take on a 13 week assignment in a beach adjacent Hospital facility in Florida. Must be ASCP certified and Florida licensed . We will pay for Housing, travel and transportation and offer a per diem. We also offer health insurance after 30 days of employment. This would be a great assignment for someone trying to escape the impending winter chill. Don Knowles Healthcare Connections Inc. 866-346-8522 ext 311 www.labcareer.com From liz <@t> premierlab.com Thu Oct 6 12:38:52 2005 From: liz <@t> premierlab.com (Elizabeth Chlipala) Date: Thu Oct 6 12:39:06 2005 Subject: [Histonet] old microtime Message-ID: <000701c5ca9c$d125ee00$a7d48a80@AMY> I know that this is a weird question to ask but I'm going to be purchasing a new LEICA RM2255 and they are currently running a special now. If I can trade in an old microtome with my new purchase I get $2,000.00 off the price of the microtome. The trade in can be any type of microtome, age does not matter, but it needs to have both a knife holder and specimen holder. If anyone out in histoland has an old non workable microtome that they are willing to part with. I'll pay for the shipping. Thanks in advance Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC P.O. Box 18592 Boulder, Colorado 80308 Office: (303) 735-5001 Fax: (303) 735-3540 liz@premierlab.com www.premierlab.com Ship to Address: Premier Laboratory University of Colorado MCDB, Room A3B40 Boulder, Colorado 80309 From GDawson <@t> dynacaremilwaukee.com Thu Oct 6 13:23:26 2005 From: GDawson <@t> dynacaremilwaukee.com (Dawson, Glen) Date: Thu Oct 6 13:28:59 2005 Subject: [Histonet] CAP question Message-ID: Sue, I've assembled a list of my primary antibodies with their corresponding clones as well as other names that might be used for each. I have about 1 billion clients so including the various processing programs each of them might have would be a monumental task so I refuse to do it. The scoring method is pathologist specific (some do not score the tests you listed at all) so the responsibility for that lies with the docs. I do the CAP immunohistochemistry survey to satisfy part of the last requirement but "evaluating interobserver variability among the pathologists" is not something I do either. I know we should all be able to do something like this in between all of our many naps & other free time activities but I just can't seem to get this vital piece of CAP legislation satisfied. There are some CAP questions, like this one, that just leave me scratching my head and wondering who the he#* is sitting around thinking this sh%@ up. I will personally try to show that I'm making an effort to satisfy some of this one and if that is not enough, I guess it'll just be a CAP ding marked up against my lab. In today's lab, one must choose their battles wisely and this battle doesn't seem to be worth the headache. Just My Opinion, Glen Dawson BS, HT & QIHC (ASCP) IHC Manager Milwaukee, WI -----Original Message----- From: Kapoor, Sue [mailto:Sue.Kapoor@uhsi.org] Sent: Thursday, October 06, 2005 7:56 AM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] CAP question Hi everyone, is anyone familiar with CAP #ANP.22925 "For IHC tests that provide independent predictive/prognostic information, does the patient report include info on specimen processing, the antibody clone, and the scoring method used? (e.g., hormone receptor in breast carcinoma, HER-2/neu, EGFR). "The lab should periodically compare its patient results with published benchmarks, and also evaluate interobserver variabitlity among the pathologists in the lab". how are others handling this?? Many, many thanks in advance, Sue Kapoor, HT (ASCP) Histology Coordinator Kenosha Medical Center Kenosha, WI 262-653-5570 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From mward <@t> wfubmc.edu Thu Oct 6 15:17:05 2005 From: mward <@t> wfubmc.edu (Martha Ward) Date: Thu Oct 6 15:17:38 2005 Subject: [Histonet] TFEB and TFE3 antibodies Message-ID: <61135F0455D33347B5AAE209B903A30410326B26@EXCHVS2.medctr.ad.wfubmc.edu> I have had a request from our Pathologist to get these antibodies and I wondered if anyone had any experience with either one of them? The papers he referenced mentioned Santa Cruz as the source. Thanks in advance for your help. Martha Ward Wake Forest University Baptist Medical Center From ploykasek <@t> phenopath.com Thu Oct 6 15:53:13 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Thu Oct 6 15:53:45 2005 Subject: [Histonet] TFEB and TFE3 antibodies In-Reply-To: <61135F0455D33347B5AAE209B903A30410326B26@EXCHVS2.medctr.ad.wfubmc.edu> Message-ID: Martha, We use the TFE3 from Santa Cruz. TFE3 is associated with a subgroup of papillary renal cell carcinomas (RCC) having a X:1 chromosome translocation. We use it with a heat pretreatment in citrate. Note that it is a goat antibody, so blocking is helpful in attaining a clean stain. Hope this helps. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information.? Any unauthorized review, use, disclosure or distribution is prohibited.? If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message or you may call PhenoPath Laboratories, Seattle, WA U.S.A at (206) 374-9000. > I have had a request from our Pathologist to get these antibodies and I > wondered if anyone had any experience with either one of them? The > papers he referenced mentioned Santa Cruz as the source. Thanks in > advance for your help. > > Martha Ward > Wake Forest University Baptist Medical Center > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From vbaker60 <@t> yahoo.com Thu Oct 6 16:42:53 2005 From: vbaker60 <@t> yahoo.com (Victoria Baker) Date: Thu Oct 6 16:43:11 2005 Subject: [Histonet] Looking for Steve Mello Message-ID: <20051006214254.16024.qmail@web52502.mail.yahoo.com> Hi! I'm looking for Steve Mello, he used to be a supervisor at Brigham & Women back around 1998. In his lab he had these instruments that were chiller stands for the blocks. They plugged in and the tech used them with ice for paraffin sections. Steve if you're out there could you get in touch with me? I'd appreciate it. Vikki Baker The Godfrey Lab Mt. Sinai School of Medicine New York, NY __________________________________ Yahoo! Mail - PC Magazine Editors' Choice 2005 http://mail.yahoo.com From AnthonyH <@t> chw.edu.au Thu Oct 6 18:19:41 2005 From: AnthonyH <@t> chw.edu.au (Tony Henwood) Date: Thu Oct 6 18:20:13 2005 Subject: [Histonet] amyloid in FNA smears Message-ID: If you are using congo red, try air-drying (you get better retention of material) followed by formalin vapor fixation. Two references that might be of use: 1. Duston, M.A.,etal (1987) Am.J.Med. 82:412-414. 2. Libbey,C.A., (1983) Arch. Intern. Med. 143;1549-1552. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Houston, Ronnie Sent: Thursday, 6 October 2005 10:14 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] amyloid in FNA smears What is the best fixation for smears from a fat-pad of a patient suspected with having amyloidosis, to get optimal preservation for demonstration of amyloid? Thanks Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com ____________________________________________________________________________ ____________________________________________________ ____________________________________________________________________________ ____________________________________________________ The information in this communication is intended to be confidential to the Individual(s) and/or Entity to whom it is addressed. It may contain information of a Privileged and/or Confidential nature, which is subject to Federal and/or State privacy regulations. In the event that you are not the intended recipient or the agent of the intended recipient, do not copy or use the information contained within this communication, or allow it to be read, copied or utilized in any manner, by any other person(s). Should this communication be received in error, please notify the sender immediately either by response e-mail or by phone, and permanently delete the original e-mail, attachment(s), and any copies. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** From AnthonyH <@t> chw.edu.au Thu Oct 6 18:21:43 2005 From: AnthonyH <@t> chw.edu.au (Tony Henwood) Date: Thu Oct 6 18:22:03 2005 Subject: [Histonet] Histology Journals Message-ID: I concur, The Journal of Histotechnology and Histologic are gems Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Rene J Buesa Sent: Thursday, 6 October 2005 11:33 PM To: Prior, Andrew; 'histonet@lists.utsouthwestern.edu' Subject: Re: [Histonet] Histology Journals Although it could be considered as a "biassed" opinion I would suggest for you to look at the Journal of Histotechnology from the National Society of Histotechnology. Rene J. "Prior, Andrew" wrote: I was wondering if anyone could recommend a good histology journal, that covers a lot of the methods and equipment. I've had a look at some journals on the web, but would appreciate some feedback from regular readers on how useful you find them. Many thanks Andrew Andrew Prior Histologist Smith & Nephew Research Centre York Science Park Heslington York, YO10 5DF UK Andrew.Prior@smith-nephew.com Confidentiality. This electronic transmission is strictly confidential to Smith & Nephew and intended solely for the addressee. It may contain information which is covered by legal, professional or other privilege. If you are not the intended addressee, or someone authorised by the intended addressee to receive transmissions on behalf of the addressee, you must not retain, disclose in any form, copy or take any action in reliance on this transmission. If you have received this transmission in error, please notify the sender as soon as possible and destroy this message. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** From Nfitzgi726 <@t> charter.net Thu Oct 6 19:39:04 2005 From: Nfitzgi726 <@t> charter.net (Noreen) Date: Thu Oct 6 19:39:30 2005 Subject: [Histonet] cryostat cleaner/decon Message-ID: <00fb01c5cad7$85948650$6400a8c0@noreen6yhmgcrn> If my memory serves me correctly last month there was a little bit of a discussion about a product that you can decon a cryostat with while it was up and running. Does anyone happen to have any information about such a thing?? Thanks, Noreen Fitzgibbons /\----/\ ( * = * ) oink From Gunilla.Aronsson <@t> odontologi.gu.se Fri Oct 7 02:30:00 2005 From: Gunilla.Aronsson <@t> odontologi.gu.se (Gunilla Aronsson) Date: Fri Oct 7 02:31:56 2005 Subject: [Histonet] storing fixed cells Message-ID: <2B6C07DD-3704-11DA-B188-000393BD4D72@odontologi.gu.se> Hi everyone, I?m planning to use cells, grown in chamber slides, to immunofluorescence. Does anyone out there know if it is possible to store these cells (human fibroblasts) for some time after fixation with ice-cold ethanol. Should they be kept in refridgerator or frozen and should they be covered by some buffer. Thanks in advance Gunilla Aronsson G?teborg University Sweden From Tanni.Ahmed <@t> intervet.com Fri Oct 7 02:40:43 2005 From: Tanni.Ahmed <@t> intervet.com (Ahmed, T (Tanni)) Date: Fri Oct 7 02:44:15 2005 Subject: [Histonet] Fixation using Modified Bouins and NBF Message-ID: <09E7875E806DFB4BBECBADBF966BDBB6F6763C@mksn79.d50.intra> John, Composition of Modified Bouins we use is - 36gms picric acid 200mls glacial acetic acid 400mls 10% NBF 3.6L Distilled water Tissues are fixed for at least 24 hrs. Bouins is changed to 70% IMS after the recommended time - i.e 24 hrs however this period can be extended up to a maximum of 3 days. We are fixing avian bursa specimens in 10% NBF at the moment and performing IHC to stain for IBD (Infectious Bursal Disease) using the Vector ABC Elite kit. Our pathologist would like to go back to the days and try using Modified Bouins to fix bursal tissues then perform the same IHC. I have never used toxic MD fixative before and trying to convince him that 10% NBF works just as well...I will have a go though! no harm in trying old techniques. Thanks, Tanni -----Original Message----- From: John A. Kiernan [mailto:jkiernan@uwo.ca] Sent: 6 October 2005 16:52 To: Ahmed, T (Tanni) Cc: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Fixation using Modified Bouins and NBF What is the composition of the "modified Bouin's" fixative, and how long did the specimens remain in it? Back in the old days Bouin's was considered a good fixative for some immunofluorescent applications, especially localizing protein and peptide hormones. The picric acid in the solution subdues much of the autofluorescence. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Ahmed, T (Tanni)" wrote: > > > Dear Histonetters, > > I would like to know if fixation of avian bursa tissue with Modified > Bouins fixative over 10% Neutral Buffered Formalin has any advantagous > effect on immuno precipitation? > > Thanks in advance, > > Tanni > > > > > > Tanni S Ahmed > > Scientific Officer - Histopathology, R&D > > > > Intervet UK Ltd. > > Walton Manor, > > Walton, Milton Keynes, > > Buckinghamshire, > > MK7 7AJ, UK. > > > > Tel. +44(0)1908 685552/685543 > > Fax +44(0)1908 685614 > > E-mail: tanni.ahmed@intervet.com > > > > > > > > -------------------------------------- > This message, including attachments, is confidential and may be > privileged. If you are not an intended recipient, please notify the > sender then delete and destroy the original message and all copies. > You should not copy, forward and/or disclose this message, in whole or > in part, without permission of the sender. > -------------------------------------- > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet -------------------------------------- This message, including attachments, is confidential and may be privileged. If you are not an intended recipient, please notify the sender then delete and destroy the original message and all copies. You should not copy, forward and/or disclose this message, in whole or in part, without permission of the sender. -------------------------------------- From BMolinari <@t> heart.thi.tmc.edu Fri Oct 7 05:41:41 2005 From: BMolinari <@t> heart.thi.tmc.edu (Molinari, Betsy) Date: Fri Oct 7 05:43:05 2005 Subject: [Histonet] Cryostats Message-ID: Hi Andrea, I use the Leica CM 1800. I use it for animal research. Mice, rabbits, dogs, pigs, and cows. We have owned it several years and have never had a problem. Betsy Molinari HT (ASCP) Texas Heart Institute Cardiovascular Pathology 6770 Bertner Ave. Houston,TX 77030 832-355-6524 832-355-6812 (fax) -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Andrea T. Hooper Sent: Thursday, October 06, 2005 10:11 AM To: Histonet Subject: [Histonet] Cryostats Hello everyone ... What is everyone's favorite cryostat and why? We are in the market. I have a few personal favorites but I wanted to hear from the general audience what is liked best. We cut mostly mouse tissue (probably 70-80% mouse tissue) and 50% is probably mouse bone. Thanks! Andrea -- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From drgrant <@t> cmh.edu Fri Oct 7 06:58:18 2005 From: drgrant <@t> cmh.edu (Grant, Debra, R) Date: Fri Oct 7 07:01:31 2005 Subject: [Histonet] cap inspection Message-ID: <6F434E27D5DE944B9E898984CADDD14904394D@exchmail.CMH.Internal> Good Morning to All, I would like some information on what other labs are keeping daily QC charts on, such as equipment in the lab? Especially whether you keep a daily QC chart on microtomes, water baths etc. This will be my first cap inspection. Thanks to all the responses on the Immuno QC sheets they were all very helpful! :-) Debby R. Grant HT(ASCP) Histology Department The Children's Mercy Hospital & Clinics 2401 Gillham Rd. Kansas City, Missouri 64108 lab (816)234-3827 fax (816) 802-1492 From bakerj <@t> umich.edu Fri Oct 7 08:02:40 2005 From: bakerj <@t> umich.edu (John Baker) Date: Fri Oct 7 08:01:34 2005 Subject: [Histonet] HISTO STAINS Message-ID: <2bdc9dafd427a1c2624d22fc40793403@umich.edu> > > Subject: HISTO STAINS > > > Dear Histonet, > > Question about staining. > > We are trying to stain paraffin embedded sections of a rat mandible in > order to > use Bioquant software to threshold mineralized bone vs. non-bone. In > the past, > we used plastic sections, and we stained them with the "Modified > Villanueva-Gomori Trichrome" for plastic. We mistakenly used this same > procedure for our paraffin sections, and we achieved the color > differentiation > we wanted as far as blue vs. red. However, the colors were not very > intense, > and we are having trouble thresholding where at the interface of bone > and > non-bone. > > Is there any stain we can use that will keep the same colors, but more > intense? > We cannot use a stain such as the "Gomori One-Step Trichrome" for > paraffin > because we cannot have any red scattered throughout the regions of > mineralized > bone (the software cannot differentiate between red that is bone and > red that > is non-bone. > > Thanks, > Krikor Arman > > > John A. Baker Histology Unit Orthopaedic Research Laboratories The University of Michigan 400 North Ingalls, G-161 Ann Arbor, MI 48103-0486 office phone:734-936-1635 From vd38 <@t> georgetown.edu Fri Oct 7 08:18:28 2005 From: vd38 <@t> georgetown.edu (Vernon Dailey) Date: Fri Oct 7 08:20:16 2005 Subject: [Histonet] Sticky Slides Message-ID: <434675A4.9070709@georgetown.edu> Hi Histonet, I am doing an in situ PCR/Hybridization. I am working with some cells that were suspended in histogel, fixed in formalin, processed and embedded. I am having some trouble keeping the tissue on the slides through all of the incubations and wash steps. I am using silanized slides and baking the slides at 60 degrees for 1 hour prior to use. Does anyone have any suggestions as to how I can get these samples to stick to the slides? Secondly, would a cytospin preparation be more likely to stay on the slide? Thanks in advance. -Vernon From anh2006 <@t> med.cornell.edu Fri Oct 7 08:20:36 2005 From: anh2006 <@t> med.cornell.edu (anh2006@med.cornell.edu) Date: Fri Oct 7 08:20:59 2005 Subject: [Histonet] storing fixed cells In-Reply-To: <2B6C07DD-3704-11DA-B188-000393BD4D72@odontologi.gu.se> References: <2B6C07DD-3704-11DA-B188-000393BD4D72@odontologi.gu.se> Message-ID: <46654.66.108.123.192.1128691236.squirrel@webmail> When I am doing staining as such, I fix in 1-4% PFA for 15 mintues and then I transfer to PBS (after at least one PBS wash). I often store in PBS for up to a week (at 4 deg C and parafilmed) before staining with no obvious change in staining quality. Andrea > Hi everyone, > > I?m planning to use cells, grown in chamber slides, to > immunofluorescence. Does anyone out there know if it is possible to > store these cells (human fibroblasts) for some time after fixation with > ice-cold ethanol. Should they be kept in refridgerator or frozen and > should they be covered by some buffer. > > Thanks in advance > > Gunilla Aronsson > > G?teborg University > Sweden > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > From anh2006 <@t> med.cornell.edu Fri Oct 7 08:25:20 2005 From: anh2006 <@t> med.cornell.edu (anh2006@med.cornell.edu) Date: Fri Oct 7 08:25:33 2005 Subject: [Histonet] Sticky Slides In-Reply-To: <434675A4.9070709@georgetown.edu> References: <434675A4.9070709@georgetown.edu> Message-ID: <46666.66.108.123.192.1128691520.squirrel@webmail> Hi Vernon, I have worked with cells prepared the way you describe and have no problems with them staying on the slide when I use Superfrost Plus slides. I cannot imagine why they are coming off on silane slides ... could it be your batch of slides that is the problem??? As far as cytospins, although I also have little trouble with cytospins coming off the slide when I use Superfrost Plus, they definitely suffer more damage during the staining process than paraffin. In my experience paraffin sections stay on better but by the sounds of it the cytospins may work better for you. Are you doing antigen retrieval, perhaps "harsh" conditions?? This may affect the integrity of the paraffin section. Obviously with the cytospin you wouldn't be doing that so that may be the way to go if your problem persists. I would look to the antigen retrieval technique and the batch of slides as the culprits first. > Hi Histonet, > > I am doing an in situ PCR/Hybridization. I am working with some cells > that were suspended in histogel, fixed in formalin, processed and > embedded. I am having some trouble keeping the tissue on the slides > through all of the incubations and wash steps. I am using silanized > slides and baking the slides at 60 degrees for 1 hour prior to use. Does > anyone have any suggestions as to how I can get these samples to stick > to the slides? Secondly, would a cytospin preparation be more likely to > stay on the slide? Thanks in advance. > > -Vernon From Nancy.Lowen <@t> med.va.gov Fri Oct 7 08:52:25 2005 From: Nancy.Lowen <@t> med.va.gov (Lowen, Nancy) Date: Fri Oct 7 08:50:00 2005 Subject: [Histonet] Type X collagen for mouse tissue Message-ID: I am looking for an antibody to mouse Type X collagen. If anyone knows who has a good one could you please let me know. Thanks in advance. Nancy.lowen@med.va.gov From dusko.trajkovic <@t> pfizer.com Fri Oct 7 09:25:17 2005 From: dusko.trajkovic <@t> pfizer.com (Trajkovic, Dusko) Date: Fri Oct 7 09:25:39 2005 Subject: [Histonet] Question about fixation Message-ID: <3AD0BD3142459B4E9B12CBEAFF2B89B2D89663@lajamrexm01.amer.pfizer.com> I received this email from a colleague. Can someone help him out???? Thank you, Dusko Trajkovic 858-638-6202 Just wanted to send you a quick summary of my question from yesterday: I wanted to know how to best fix late term mouse embryo heads, such as e14.5 and onward, esp injected with a virus that expresses GFP. What I would ideally like to do is to cut, mount, stain with DAPI, and directly visualize by fluorescence microscopy. Immersion-fixation in 4% PFA does not seem to work that well in general for head fixation. Would formalin immersion-fixation work better? Or other alternative fixatives? What about autofluorescence from these other fixatives? Thanks and see you later, Yun Yun C. Yung Graduate student Chun Lab, Dept of Molecular Biology The Scripps Research Institute yyung@scripps.edu www.scripps.edu/mb/chun ---------------------------------------------------------------------- LEGAL NOTICE Unless expressly stated otherwise, this message is confidential and may be privileged. It is intended for the addressee(s) only. Access to this E-mail by anyone else is unauthorized. If you are not an addressee, any disclosure or copying of the contents of this E-mail or any action taken (or not taken) in reliance on it is unauthorized and may be unlawful. If you are not an addressee, please inform the sender immediately. From Janet.Bonner <@t> FLHosp.org Fri Oct 7 09:56:15 2005 From: Janet.Bonner <@t> FLHosp.org (Bonner, Janet) Date: Fri Oct 7 09:56:39 2005 Subject: [Histonet] Cryostats Message-ID: <07AB60D5D7B9754EBF56F360F98D083DEB442F@fh2k093.fhmis.net> I second that!! We have the Leica 1800 and 1900's- eight of them - we're a large hospital system of seven hospitals and lots of frozens all day long. Rarely have we had a problem with them. Janet, Florida Hospital Orlando -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Molinari, Betsy Sent: Friday, October 07, 2005 6:42 AM To: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Cryostats Hi Andrea, I use the Leica CM 1800. I use it for animal research. Mice, rabbits, dogs, pigs, and cows. We have owned it several years and have never had a problem. Betsy Molinari HT (ASCP) Texas Heart Institute Cardiovascular Pathology 6770 Bertner Ave. Houston,TX 77030 832-355-6524 832-355-6812 (fax) -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Andrea T. Hooper Sent: Thursday, October 06, 2005 10:11 AM To: Histonet Subject: [Histonet] Cryostats Hello everyone ... What is everyone's favorite cryostat and why? We are in the market. I have a few personal favorites but I wanted to hear from the general audience what is liked best. We cut mostly mouse tissue (probably 70-80% mouse tissue) and 50% is probably mouse bone. Thanks! Andrea -- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From denise <@t> pclv.net Fri Oct 7 10:16:50 2005 From: denise <@t> pclv.net (Denise) Date: Fri Oct 7 10:15:28 2005 Subject: [Histonet] Histology Gross Tech Message-ID: Phoenix Central Laboratory for Veterinarians, Inc. has a position available for an HT or eligible person to perform gross dissection of veterinary tissue specimens. This is a part-time weekend position. Please submit your resume to Denise DeRouchey, c/o Phoenix Central Laboratory, 11620 Airport Road, Everett, WA 98204. Alternately, you may fax your resume to 425-355-3676. Phoenix Central Laboratory, (PCLV), is a veterinary reference laboratory that provides diagnostic services to more than 800 clinics located throughout the Pacific Northwest and Alaska. We are dedicated to providing rapid, quality service. Our laboratory is located in Everett, Washington, just thirty minutes north of downtown Seattle. From tpmorken <@t> labvision.com Fri Oct 7 10:44:43 2005 From: tpmorken <@t> labvision.com (Morken, Tim - Labvision) Date: Fri Oct 7 10:45:07 2005 Subject: [Histonet] Cryostats Message-ID: <0556BE8AC5551E4E8AF6BB9E42509BA20328D1DA@usca0082k08.labvision.apogent.com> We have a Microm (Richard Allan) 550 with a built-in vacuum - it never gets dirty, and the vacuum can actually be used instead of a brush to straighten out sections. Tim Morken Lab Vision - Neomarkers www.labvision.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Molinari, Betsy Sent: Friday, October 07, 2005 3:42 AM To: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Cryostats Hi Andrea, I use the Leica CM 1800. I use it for animal research. Mice, rabbits, dogs, pigs, and cows. We have owned it several years and have never had a problem. Betsy Molinari HT (ASCP) Texas Heart Institute Cardiovascular Pathology 6770 Bertner Ave. Houston,TX 77030 832-355-6524 832-355-6812 (fax) -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Andrea T. Hooper Sent: Thursday, October 06, 2005 10:11 AM To: Histonet Subject: [Histonet] Cryostats Hello everyone ... What is everyone's favorite cryostat and why? We are in the market. I have a few personal favorites but I wanted to hear from the general audience what is liked best. We cut mostly mouse tissue (probably 70-80% mouse tissue) and 50% is probably mouse bone. Thanks! Andrea -- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jkiernan <@t> uwo.ca Fri Oct 7 12:02:38 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Fri Oct 7 12:03:05 2005 Subject: [Histonet] Fixation using Modified Bouins and NBF References: <09E7875E806DFB4BBECBADBF966BDBB6F6763C@mksn79.d50.intra> Message-ID: <4346AA2E.FDDDFD38@uwo.ca> The modified Bouin that you describe is an illogical mixture. The purpose of Bouin's fixative is to obtain the advantages of acetic acid (good nuclear morphology) and picric acid (rapid coagulation of cytoplasm and strong staining by anionic dyes) in addition to protein cross-linking bt formaldehyde. Your modification uses neutral buffered formaldehyde, which will oppose the required acidity. The correct recipe for Bouin's fixative is: Saturated aqueous picric acid 750 ml Formalin (37-40% HCHO) 250 ml Acetic acid (glacial) 50 ml Fix usually for 24 hours, then transfer to 70% alcohol. If the tissue is still yellow in the paraffin block, put the hydrated sections in something weakly alkaline to remove residual picric acid, then was well with water before staining. Retained picric acid interferes with some stains, including those used for blood cells. If red blood cells are of interest, don't use Bouin's fluid for fixation because it damages them; use neutral, buffered formaldehyde instead. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Ahmed, T (Tanni)" wrote: > > John, > > Composition of Modified Bouins we use is - > 36gms picric acid > 200mls glacial acetic acid > 400mls 10% NBF > 3.6L Distilled water > > Tissues are fixed for at least 24 hrs. Bouins is changed to 70% IMS > after the recommended time - i.e 24 hrs however this period can be > extended up to a maximum of 3 days. > > We are fixing avian bursa specimens in 10% NBF at the moment and > performing IHC to stain for IBD (Infectious Bursal Disease) using the > Vector ABC Elite kit. Our pathologist would like to go back to the days > and try using Modified Bouins to fix bursal tissues then perform the > same IHC. I have never used toxic MD fixative before and trying to > convince him that 10% NBF works just as well...I will have a go though! > no harm in trying old techniques. > > Thanks, > > Tanni > > -----Original Message----- > From: John A. Kiernan [mailto:jkiernan@uwo.ca] > Sent: 6 October 2005 16:52 > To: Ahmed, T (Tanni) > Cc: histonet@lists.utsouthwestern.edu > Subject: Re: [Histonet] Fixation using Modified Bouins and NBF > > What is the composition of the "modified Bouin's" > fixative, and how long did the specimens remain in > it? Back in the old days Bouin's was considered a > good fixative for some immunofluorescent applications, especially > localizing protein and peptide hormones. The picric acid in the solution > subdues much of the autofluorescence. > -- > ------------------------------- > John A. Kiernan > Department of Anatomy and Cell Biology > The University of Western Ontario > London, Canada N6A 5C1 > kiernan[AT]uwo.ca > http://publish.uwo.ca/~jkiernan/ > http://instruct.uwo.ca/anatomy/530/index.htm > _______________________________ > "Ahmed, T (Tanni)" wrote: > > > > > > Dear Histonetters, > > > > I would like to know if fixation of avian bursa tissue with Modified > > Bouins fixative over 10% Neutral Buffered Formalin has any advantagous > > > effect on immuno precipitation? > > > > Thanks in advance, > > > > Tanni > > > > > > > > > > > > Tanni S Ahmed > > > > Scientific Officer - Histopathology, R&D > > > > > > > > Intervet UK Ltd. > > > > Walton Manor, > > > > Walton, Milton Keynes, > > > > Buckinghamshire, > > > > MK7 7AJ, UK. > > > > > > > > Tel. +44(0)1908 685552/685543 > > > > Fax +44(0)1908 685614 > > > > E-mail: tanni.ahmed@intervet.com > > From settembr <@t> umdnj.edu Fri Oct 7 11:44:24 2005 From: settembr <@t> umdnj.edu (Dana Settembre) Date: Fri Oct 7 12:04:49 2005 Subject: [Histonet] cap inspection Message-ID: Debby, We keep a daily QC chart on the water baths for temperature and daily QC chart on the water baths and microtomes for daily maintenance such as cleaning. Dana Settembre University Hospital >>> "Grant, Debra, R" 10/7/2005 7:58:18 AM >>> Good Morning to All, I would like some information on what other labs are keeping daily QC charts on, such as equipment in the lab? Especially whether you keep a daily QC chart on microtomes, water baths etc. This will be my first cap inspection. Thanks to all the responses on the Immuno QC sheets they were all very helpful! :-) Debby R. Grant HT(ASCP) Histology Department The Children's Mercy Hospital & Clinics 2401 Gillham Rd. Kansas City, Missouri 64108 lab (816)234-3827 fax (816) 802-1492 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From PIXLEYSK <@t> UCMAIL.UC.EDU Fri Oct 7 12:27:56 2005 From: PIXLEYSK <@t> UCMAIL.UC.EDU (Pixley, Sarah (pixleysk)) Date: Fri Oct 7 12:28:11 2005 Subject: [Histonet] RE: labelling cells in culture Message-ID: Dear Liz: The compound that is designed for that type of experiment is called Cell Trakker Green, from Molecular Probes. You incubate that with your living cells (30 mins, incubator), then they are labeled with a fluorescent compound that is green, non-toxic and that stays within the cells for their lifespan. It also stays within the cells after fixation. (fluorescein optics on your microscope) I have labeled cells and then transferred them to another co-culture and kept them going for one week before fixing and I was able to still detect labeled cells. I am not sure about how well they will last for 3 weeks. But it was the best thing I could find for the type of thing you are describing. It is also used for transplantation experiments. Good luck! Sarah Univ. Cincinnati From adelsyscarol <@t> yahoo.com Fri Oct 7 13:26:32 2005 From: adelsyscarol <@t> yahoo.com (Carol Wilson) Date: Fri Oct 7 13:26:51 2005 Subject: [Histonet] Looking for Cytoclips and Surgipath VCP Cassette Labeler Message-ID: <20051007182632.12054.qmail@web51512.mail.yahoo.com> Anyone in histoland/cytoland that has gone to using disposable clips for their Thermo/Shandon cytospin and is interested in getting rid of their Cytoclips, please contact me. And since I'm asking, if anyone has a Surgipath VCP cassette labeler sitting around collecting dust and would be interested in selling, contact me. Thanks, Carol Carol Wilson, HT(ASCP) Histology and Laboratory Products and Service Adelsys,Inc. 1925 Lee Rd. Cleveland Heights, OH 44118 216-932-7500 Fax 216-932-7850 --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. From hymclab <@t> hyhc.com Fri Oct 7 13:40:07 2005 From: hymclab <@t> hyhc.com (hymclab) Date: Fri Oct 7 13:31:54 2005 Subject: [Histonet] cap inspection Message-ID: Debby, We also keep daily QC charts on our waterbath temps and cleaning of mircrotomes. In addition, daily, we record temps of embedding center, cryostat (also, cleaning of cryostat), refrigerators, room temperature, processors (along with rotating of solutions). We also keep record of when solutions are changed and rotated in our H&E line and our frozen line. Then of course there are the charts for recording special staines and immunos. Hope this helps, Dawn -----Original Message----- From: Dana Settembre [mailto:settembr@umdnj.edu] Sent: Friday, October 07, 2005 11:44 AM To: drgrant@cmh.edu; histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] cap inspection Debby, We keep a daily QC chart on the water baths for temperature and daily QC chart on the water baths and microtomes for daily maintenance such as cleaning. Dana Settembre University Hospital >>> "Grant, Debra, R" 10/7/2005 7:58:18 AM >>> Good Morning to All, I would like some information on what other labs are keeping daily QC charts on, such as equipment in the lab? Especially whether you keep a daily QC chart on microtomes, water baths etc. This will be my first cap inspection. Thanks to all the responses on the Immuno QC sheets they were all very helpful! :-) Debby R. Grant HT(ASCP) Histology Department The Children's Mercy Hospital & Clinics 2401 Gillham Rd. Kansas City, Missouri 64108 lab (816)234-3827 fax (816) 802-1492 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From ploykasek <@t> phenopath.com Fri Oct 7 13:54:27 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Fri Oct 7 13:55:03 2005 Subject: [Histonet] Zymed stainer Message-ID: Has anyone tried the immunostainer from Zymed - "Mozaic"? I would appreciate any feedback on ease of use, quality of staining, etc... Thanks for the input. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information.? Any unauthorized review, use, disclosure or distribution is prohibited.? If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message or you may call PhenoPath Laboratories, Seattle, WA U.S.A at (206) 374-9000. From gintini <@t> buffalo.edu Fri Oct 7 15:22:26 2005 From: gintini <@t> buffalo.edu (Beppe Intini) Date: Fri Oct 7 15:22:39 2005 Subject: [Histonet] Bone tissue processing Message-ID: Dear Histonetters, I am a PhD student in Oral Biology at the University at Buffalo, NY - USA. I am "desperately" looking for a lab/facility where i can send some of my rat calvaria bone samples for undecalcified tissue embedding/processing/staining. Unfortunately at the University at Buffalo the Histo/Pathology facility processes only decalcified bone. Can anyone help? Is there any facility out there that can process my bone samples for a relatively reasonable price? Thank you. Beppe Intini -- Giuseppe Intini Clinical Assistant Professor - Periodontics PhD Candidate - Oral Biology University at Buffalo Buffalo, NY 14214 From godsgirlnow <@t> msn.com Fri Oct 7 15:57:26 2005 From: godsgirlnow <@t> msn.com (Roxanne Soto) Date: Fri Oct 7 15:57:46 2005 Subject: [Histonet] VENDOR HELP Message-ID: I am in the need of some capital equipment for urine cytology. We do lots of urine cytology and I need a cytofuge that holds more than 4 slides and a centrafuge that can spin down more tha 4 of the 50 ml conical tubes at a time as well---preferrably both will do the same amount. Please let me know if you can help ASAP. I will be needing 2 of each. Thanks Roxanne Soto Tampa From Delventh3 <@t> aol.com Sat Oct 8 08:43:36 2005 From: Delventh3 <@t> aol.com (Delventh3@aol.com) Date: Sat Oct 8 08:43:54 2005 Subject: [Histonet] Re: Histonet Digest, Vol 23, Issue 9 Message-ID: <1ac.4143e951.30792708@aol.com> Hello Don - I have my Florida license. I am retired from over 40 years as a histotechnologist. For the past 2 years I have been working temporary assignments. Depending upon the needs of the opening you have I might be interested in being considered for this assignment. My telephone number is 210-375-4622. If you receive the answering machine, I have my cell phone, 307-851-5595. I have attached my resume. Priscilla Delventhal BA, HT, HTL (ASCP) -------------- next part -------------- Pris 3900 Habersham Schertz, Texas Voice: 210-375-4622  Cel E-Ma Revised: May, 2005 EDUCATION: BA - Thomas A. Edison State College, Trenton, New Jersey HT, HTL (ASCP) PROFESSIONAL ORGANIZATIONS: National Society of Histotechnologists American Society of Clinical Pathologists AWARDS: 1982 Who's Who in American Women STAT Flor WORK EXPERIENCE June,1961 –July, 1964 Assistant supervisor, Surgical Pathology Laboratory,          Â Â Â Â Â  University of Colorado Medical School, Denver, Colorado - some research in tissue culture. July Cytology slides. Denv Sept Hospital, Wheatridge, Colorado.  General Histologic technic.  Cytology preparation and screening. September,1965 – December,1968 Assistant supervisior, Histolog Veterinary Medicine, Ft. Collins, Colorado. General Histo Hematology research. In December of 1968 we moved to Wyoming for my husband's job. There we September,1970- May,1971 Filled in for Elementary/Jr. High Music Teach May, absence of a patholo September,1973- May,1984 Histology Section Head, Pathology Laboratory Associates, Lander, Wyoming. This position included limited grossing o May, 1984 – Pathology Laboratory Assiciates closed.  No work for three months. Augu Valley Medical Center, Lander, Wyomi limited grossing of specimens and extensive cytopreparation. I left this position because an Anatomic Lab was star I lived. August,1989 – August, 2001 Histology/Cytology section head, Ri routine Histology, this position included extensive cytopreparation. I left th was retired) and I wanted do temporary work so I could see some of the vast country that we liv have not regretted this decision. August, 2001 Temporary histology jobs Univ Supervisor, Nancy at 303-316-9240, September 3, 2001 to November 2, 2001. St. Anthony's Hospital, Rockford, Illinois, Supervisor Leta Stewart at 815-395-5109. December 3, 2001 to February, 2002 Southeastern Pathology, Rome,Ga, small 3 week Histology assignment in March 2002. Telephone number 706-291-8702. Lakes Regional Hospital, Laconia, New Hampshire, Pathology Department. number 603-524-3211 ext. 3230. Returned to St. Anthony's Hospital, Rockford, Illinois, June through Septemb November 2002 to August 2003, Riverton Memorial Hospital, Riverton, Wyoming. Histology/Cytology section head. Riverton Memorial was unsuccessful in recruiting a certified histotechnologist after I left in the Summer of 2001. I returned for a year to help train 2 people in simple histology technic. This position will was finished in August and I returned to temporary assignments at that time. November, 2003 - a small 3 week assignment at Mountain View Medical Center in Los Cruces, New Mexico to cover for the sing histology technologist to have a vacation. March - July, 2004, I returned to St. Anthony's Hospital in Rockford, for another extended assignment. All routine and special Histology, plus extensive Cytology preparations. July, 2004, preparing for a move to the San Antonio, Texas area.       August, 20 2005  Pro Path Labs in Dallas, Texas.  This was mainly cutting histology            August 3 Gables Hospital, Coral Gables, Florida.  This is a single tech lab.  I did a stains, coverslipping, frozen sectio while she went on vacation.  Everything in this lab is done man ually except for tissue processing. I will return in December, 2005 for another we   Most of my permanent positions have been as section head or in a supervisory capacity. I have set up 2 laboratories, both included Histology and Cytology departments. This included purchasing equipment, setting up CPT codes and revenue codes, writing manuals and keepi I have phlebotomy experience and limited clinical laboratory experience. Computer systems I have used include the following: Co-Path Cerner Meditech I have used several different brands of automated processors, coverslippers Microm. I have also use stains and am familiar with the Ventanna Nexus autostainer for IHC.  very limited, but I am comfortable doing special stains with it. I am also very comfortable doing everything manually. In addition to all routine Histology and Cytopreparatory procedures, I have stained and screened the trichromes for 0&P's, assisted with bone marrow processed Semen analysis specimens. I have also contracted with an independent transcriptionist and coordinated her activities and kept those reports organized and delivered to all necessary departments and physicians. < Several years ago I tried to set up immunohistochemistry staining in 2 different laboratories in Wyoming, but because of the small volume these labs had, it was decided to send these tes reference laboratory. I was one of the first in Wyoming to use th Microwave oven to speed up Histologic Special Stains and Frozen Sections. I have beginning Immunofluorescence experience. I am a DOT drug collector (both urine and breathalyzer). I am computer literate. I have worked with the Wyoming State tumor registry and find it very interesting. I have assisted with Clinicians and Pathologists in researching topics for CME's and preparing the visual aids for those projects. My strengths are general histology, special stains, ribbon sections, Cryosta many years of cytoprepratory experience and a few years of cytology screening. I find great satisfaction in teaching the skill and theory of Histology. My laboratories are efficiently run, both time and monetary wise. Priscilla J. Delventhal References available upon request You may contact any of the places that I have worked including the temporary assignments. &nbs &nbs From AnthonyH <@t> chw.edu.au Sun Oct 9 18:35:15 2005 From: AnthonyH <@t> chw.edu.au (Tony Henwood) Date: Sun Oct 9 18:35:53 2005 Subject: [Histonet] Fixation using Modified Bouins and NBF Message-ID: Why use NBF (which I assume is neutral buffered formalin) when there are two acids present? Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Ahmed, T (Tanni) Sent: Friday, 7 October 2005 5:41 PM To: John A. Kiernan Cc: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Fixation using Modified Bouins and NBF John, Composition of Modified Bouins we use is - 36gms picric acid 200mls glacial acetic acid 400mls 10% NBF 3.6L Distilled water Tissues are fixed for at least 24 hrs. Bouins is changed to 70% IMS after the recommended time - i.e 24 hrs however this period can be extended up to a maximum of 3 days. We are fixing avian bursa specimens in 10% NBF at the moment and performing IHC to stain for IBD (Infectious Bursal Disease) using the Vector ABC Elite kit. Our pathologist would like to go back to the days and try using Modified Bouins to fix bursal tissues then perform the same IHC. I have never used toxic MD fixative before and trying to convince him that 10% NBF works just as well...I will have a go though! no harm in trying old techniques. Thanks, Tanni -----Original Message----- From: John A. Kiernan [mailto:jkiernan@uwo.ca] Sent: 6 October 2005 16:52 To: Ahmed, T (Tanni) Cc: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Fixation using Modified Bouins and NBF What is the composition of the "modified Bouin's" fixative, and how long did the specimens remain in it? Back in the old days Bouin's was considered a good fixative for some immunofluorescent applications, especially localizing protein and peptide hormones. The picric acid in the solution subdues much of the autofluorescence. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Ahmed, T (Tanni)" wrote: > > > Dear Histonetters, > > I would like to know if fixation of avian bursa tissue with Modified > Bouins fixative over 10% Neutral Buffered Formalin has any advantagous > effect on immuno precipitation? > > Thanks in advance, > > Tanni > > > > > > Tanni S Ahmed > > Scientific Officer - Histopathology, R&D > > > > Intervet UK Ltd. > > Walton Manor, > > Walton, Milton Keynes, > > Buckinghamshire, > > MK7 7AJ, UK. > > > > Tel. +44(0)1908 685552/685543 > > Fax +44(0)1908 685614 > > E-mail: tanni.ahmed@intervet.com > > > > > > > > -------------------------------------- > This message, including attachments, is confidential and may be > privileged. If you are not an intended recipient, please notify the > sender then delete and destroy the original message and all copies. > You should not copy, forward and/or disclose this message, in whole or > in part, without permission of the sender. > -------------------------------------- > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet -------------------------------------- This message, including attachments, is confidential and may be privileged. If you are not an intended recipient, please notify the sender then delete and destroy the original message and all copies. You should not copy, forward and/or disclose this message, in whole or in part, without permission of the sender. -------------------------------------- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From AnthonyH <@t> chw.edu.au Sun Oct 9 19:01:08 2005 From: AnthonyH <@t> chw.edu.au (Tony Henwood) Date: Sun Oct 9 19:01:37 2005 Subject: [Histonet] Fixation using Modified Bouins and NBF Message-ID: Sorry John, I just read your response. Said a lot better than I!!! Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Tony Henwood Sent: Monday, 10 October 2005 9:35 AM To: 'Ahmed, T (Tanni)'; John A. Kiernan Cc: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Fixation using Modified Bouins and NBF Why use NBF (which I assume is neutral buffered formalin) when there are two acids present? Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Ahmed, T (Tanni) Sent: Friday, 7 October 2005 5:41 PM To: John A. Kiernan Cc: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Fixation using Modified Bouins and NBF John, Composition of Modified Bouins we use is - 36gms picric acid 200mls glacial acetic acid 400mls 10% NBF 3.6L Distilled water Tissues are fixed for at least 24 hrs. Bouins is changed to 70% IMS after the recommended time - i.e 24 hrs however this period can be extended up to a maximum of 3 days. We are fixing avian bursa specimens in 10% NBF at the moment and performing IHC to stain for IBD (Infectious Bursal Disease) using the Vector ABC Elite kit. Our pathologist would like to go back to the days and try using Modified Bouins to fix bursal tissues then perform the same IHC. I have never used toxic MD fixative before and trying to convince him that 10% NBF works just as well...I will have a go though! no harm in trying old techniques. Thanks, Tanni -----Original Message----- From: John A. Kiernan [mailto:jkiernan@uwo.ca] Sent: 6 October 2005 16:52 To: Ahmed, T (Tanni) Cc: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Fixation using Modified Bouins and NBF What is the composition of the "modified Bouin's" fixative, and how long did the specimens remain in it? Back in the old days Bouin's was considered a good fixative for some immunofluorescent applications, especially localizing protein and peptide hormones. The picric acid in the solution subdues much of the autofluorescence. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Ahmed, T (Tanni)" wrote: > > > Dear Histonetters, > > I would like to know if fixation of avian bursa tissue with Modified > Bouins fixative over 10% Neutral Buffered Formalin has any advantagous > effect on immuno precipitation? > > Thanks in advance, > > Tanni > > > > > > Tanni S Ahmed > > Scientific Officer - Histopathology, R&D > > > > Intervet UK Ltd. > > Walton Manor, > > Walton, Milton Keynes, > > Buckinghamshire, > > MK7 7AJ, UK. > > > > Tel. +44(0)1908 685552/685543 > > Fax +44(0)1908 685614 > > E-mail: tanni.ahmed@intervet.com > > > > > > > > -------------------------------------- > This message, including attachments, is confidential and may be > privileged. If you are not an intended recipient, please notify the > sender then delete and destroy the original message and all copies. > You should not copy, forward and/or disclose this message, in whole or > in part, without permission of the sender. > -------------------------------------- > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet -------------------------------------- This message, including attachments, is confidential and may be privileged. If you are not an intended recipient, please notify the sender then delete and destroy the original message and all copies. You should not copy, forward and/or disclose this message, in whole or in part, without permission of the sender. -------------------------------------- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From gillian.2.brown <@t> gsk.com Mon Oct 10 03:07:01 2005 From: gillian.2.brown <@t> gsk.com (gillian.2.brown@gsk.com) Date: Mon Oct 10 03:07:38 2005 Subject: [Histonet] Metachromasia of mast cells in epoxy resin sections In-Reply-To: <4346B66E.E34CD1D7@uwo.ca> Message-ID: John, could you give me your thoughts then on the methods used in Interleukin-2?Inducible T Cell Kinase Regulates Mast Cell Degranulation and Acute Allergic Responses Johan Forssell et al Am J Respir Cell Mol Biol Vol 32. pp 511?520, 2005, in which they used and cite the methodology of Dvorak AM, Tepper RI, Weller PF, Morgan ES, Estrella P, Monahan ER, Galli SJ. Piecemeal degranulation of mast cells in the inflammatory eyelid lesions of interleukin-4 transgenic mice: evidence of mast cell histamine release in vivo by diamine oxidase-gold enzyme-affinity ultrastructural cytochemistry. Blood 1994;83:3600?3612. and Oliani SM, Lim LH, Christian HC, Pell K, Das AM, Perretti M. Morphological alteration of peritoneal mast cells and macrophages in the mouse peritoneal cavity during the early phases of an allergic inflammatory reaction. Cell Biol Int 2001;25:795?803. Am I on a hiding to nothing? Regards Gill Target Localisation Target Discovery ri- CEDD. GlaxoSmithKline Medicines Research Centre, Gunnelswood Road, STEVENAGE, Hertfordshire. SG1 2NY tel. +44 (0)1438 764119 fax. +44 (0)1438 764782 email. gillian.2.brown@gsk.com http://ukdiscovery.gsk.com/histopathology/default.htm "John A. Kiernan" 07-Oct-2005 18:54 To gillian.2.brown@gsk.com cc Subject Re: [Histonet] Metachromasia of mast cells in epoxy resin sections I find it hard to believe that the blue or red staining of mast cell granules, in osmicated and plastic-embedded tissue, would have any phsiological significance. I've counted great numbers of degranulating mast cells in paraffin sections of rat skin and respiratory passages, after fixation in Carnoy, SUSA and NBF. The granules were metachromatic (toluidine blue pH 4) inside intact mast cells and after extrusion from the cells. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ gillian.2.brown@gsk.com wrote: > > Hi Histonetters, > > would any one have experience of counting granules in mouse mast cells in > tissues which have been formaldehyde fixed, decalcified in a formic acid > based solution, post osmicated and embedded in epoxy resin? I'm taking 1 > micron sections and looking to assess the 'degranulation' status of each > mast cell in terms of how many are still dark blue and how many are pink > ie based on the metachromatic properties of toluidine blue. I suppose my > question is will the formic acid have altered the dye binding properties > so that I'll get erroneous results? I'm going to experiment with tissue > that does not normally have this step but any insights now would be most > welcome. > > Many thanks > Gill Brown > > GlaxoSmithKline Medicines Research Centre, > STEVENAGE, > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From sluhisto <@t> yahoo.com Mon Oct 10 09:24:51 2005 From: sluhisto <@t> yahoo.com (Histology SLU) Date: Mon Oct 10 09:25:09 2005 Subject: [Histonet] CAP Inspection +++_- - - - Message-ID: <20051010142451.988.qmail@web51003.mail.yahoo.com> Hello All: I am due for a CAP inspection later this month. Would any of you mind sharing your last CAP experiences both positive and negative? Thanks so much. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Mon Oct 10 09:43:24 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Mon Oct 10 09:43:36 2005 Subject: [Histonet] CAP Inspection +++_- - - - In-Reply-To: <20051010142451.988.qmail@web51003.mail.yahoo.com> Message-ID: <20051010144324.74142.qmail@web61216.mail.yahoo.com> The outcome can vary depending on how well run your laboratory is and who is doing the inspection (and how detailed oriented they are). You have to be sure that fundamental aspects of your lab are up to date, and usually a small non-compliant aspect could be expected. The most important thing is that anything that is found that needs to be corrected is both explained and corrected. Safety is important and anything that is a regulation that has to do directly with patient care is OK. Cofidentiality issues are important, as any paper-trail on diagnosis and the kind. Take your time to go, CAP list in hand, reviewing all the headings, and if you find one that is in no compliance and cannot be taken care off before the inspection, you should be the one to point it out and not wait for the inspectors to find it out. Come "always clean" with any problem. Relax, take a deep breath, be calm and everything will be OK. Rene J. Histology SLU wrote: Hello All: I am due for a CAP inspection later this month. Would any of you mind sharing your last CAP experiences both positive and negative? Thanks so much. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From sjchtascp <@t> yahoo.com Mon Oct 10 09:59:20 2005 From: sjchtascp <@t> yahoo.com (Steven Coakley) Date: Mon Oct 10 09:59:40 2005 Subject: [Histonet] deep metal embedding molds Message-ID: <20051010145920.33515.qmail@web90203.mail.scd.yahoo.com> Good morning, I'd like to pick up a couple deeper base molds for those occasional tissues requirering such. I'f anyone has a couple 12mm deep metal base molds no larger then 31mm X 23mm that they no longer use I'd appreciate hearing from you. Thanks, Steve --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Mon Oct 10 10:03:14 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Mon Oct 10 10:03:27 2005 Subject: [Histonet] CAP question In-Reply-To: Message-ID: <20051010150314.20970.qmail@web61215.mail.yahoo.com> I never fought this battle because it is outside the "realm of the histology supervisor/manager", it is entirely within the "realm of the head pathologist"; reporting and their contents is their responsibility. I always allowed them to fight this battle, and the outcome was theirs. CAP inspectors are pathologists, let them work this out amongst themselves. I did that and never got any problem. I shouwed them the regulations I could not take care of and they would take care of them. Pathologists know how to talk amongst themselves! Rene J. "Dawson, Glen" wrote: Sue, I've assembled a list of my primary antibodies with their corresponding clones as well as other names that might be used for each. I have about 1 billion clients so including the various processing programs each of them might have would be a monumental task so I refuse to do it. The scoring method is pathologist specific (some do not score the tests you listed at all) so the responsibility for that lies with the docs. I do the CAP immunohistochemistry survey to satisfy part of the last requirement but "evaluating interobserver variability among the pathologists" is not something I do either. I know we should all be able to do something like this in between all of our many naps & other free time activities but I just can't seem to get this vital piece of CAP legislation satisfied. There are some CAP questions, like this one, that just leave me scratching my head and wondering who the he#* is sitting around thinking this sh%@ up. I will personally try to show that I'm making an effort to satisfy some of this one and if that is not enough, I guess it'll just be a CAP ding marked up against my lab. In today's lab, one must choose their battles wisely and this battle doesn't seem to be worth the headache. Just My Opinion, Glen Dawson BS, HT & QIHC (ASCP) IHC Manager Milwaukee, WI -----Original Message----- From: Kapoor, Sue [mailto:Sue.Kapoor@uhsi.org] Sent: Thursday, October 06, 2005 7:56 AM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] CAP question Hi everyone, is anyone familiar with CAP #ANP.22925 "For IHC tests that provide independent predictive/prognostic information, does the patient report include info on specimen processing, the antibody clone, and the scoring method used? (e.g., hormone receptor in breast carcinoma, HER-2/neu, EGFR). "The lab should periodically compare its patient results with published benchmarks, and also evaluate interobserver variabitlity among the pathologists in the lab". how are others handling this?? Many, many thanks in advance, Sue Kapoor, HT (ASCP) Histology Coordinator Kenosha Medical Center Kenosha, WI 262-653-5570 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From jefthompson <@t> salud.unm.edu Mon Oct 10 10:53:26 2005 From: jefthompson <@t> salud.unm.edu (Jeffrey Thompson) Date: Mon Oct 10 10:54:06 2005 Subject: [Histonet] microwave antigen retrieval Message-ID: Dear Histonetters, Can anyone recommend a good microwave for antigen retrieval? The cheap oven we have gives inconsistent temperatures that I don't want to rely on any longer. I haven't been able to find an adjustable wattage model that is not extremely high end, but I don't know if the adjustability is desirable but not absolutely essential. Any advice would be appreciated. I apologize if this is a hackneyed question and I haven't kept up. Thanks, Jeff Thompson From rjbuesa <@t> yahoo.com Mon Oct 10 11:18:01 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Mon Oct 10 11:18:11 2005 Subject: [Histonet] microwave antigen retrieval In-Reply-To: Message-ID: <20051010161801.24308.qmail@web61218.mail.yahoo.com> First of all I don't know why you want to do HIER with a microwave oven (MWO). I used the MWO to heat the solutions (expedite the process) before doing the actual HIER in a steamer (any regular steamer could do) I used a Black & Decker (if it happens that you perform Her 2 Neu in your lab, DakoCytomation may provide you with the steamer for free). In the MWO you cannot obtain above 100 Celsius, and in the steamer you will get the same. If you want to do HIER above 100 celsius you will have to use a pressure cooker. Anyway, I don't see any advantage to do HIER with a MWO. I would suggest you to use a steamer (cheaper, more consistent and widely used). A good reliable MWO with wattage control will cost you many many times more than a steamer. If you insist in using the MWO at least you will have to calibrate it. I recommed you to read Journal of Histotechnology 25(1):39-43 (2002). Rene J. Jeffrey Thompson wrote: Dear Histonetters, Can anyone recommend a good microwave for antigen retrieval? The cheap oven we have gives inconsistent temperatures that I don't want to rely on any longer. I haven't been able to find an adjustable wattage model that is not extremely high end, but I don't know if the adjustability is desirable but not absolutely essential. Any advice would be appreciated. I apologize if this is a hackneyed question and I haven't kept up. Thanks, Jeff Thompson _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From Jackie.O'Connor <@t> abbott.com Mon Oct 10 11:48:26 2005 From: Jackie.O'Connor <@t> abbott.com (Jackie M O'Connor) Date: Mon Oct 10 11:48:59 2005 Subject: [Histonet] microwave antigen retrieval Message-ID: Not every lab can afford the steamer. Not every lab does Her2. Microwaves work just fine if you pay attention. As far as microwaves not reaching above 100C - I guess I'd better quit boiling water in mine at home - it could likely explode or something. My microwave in the lab went well over 100C when it melted my glass slides in a plastic coplin jar . . . .I have photos. I think Sigma still has a good lab quality microwave. Rene J Buesa Sent by: histonet-bounces@lists.utsouthwestern.edu 10/10/2005 11:18 AM To: Jeffrey Thompson , histonet@lists.utsouthwestern.edu cc: (bcc: Jackie M O'Connor/LAKE/GPRD/ABBOTT) Subject: Re: [Histonet] microwave antigen retrieval First of all I don't know why you want to do HIER with a microwave oven (MWO). I used the MWO to heat the solutions (expedite the process) before doing the actual HIER in a steamer (any regular steamer could do) I used a Black & Decker (if it happens that you perform Her 2 Neu in your lab, DakoCytomation may provide you with the steamer for free). In the MWO you cannot obtain above 100 Celsius, and in the steamer you will get the same. If you want to do HIER above 100 celsius you will have to use a pressure cooker. Anyway, I don't see any advantage to do HIER with a MWO. I would suggest you to use a steamer (cheaper, more consistent and widely used). A good reliable MWO with wattage control will cost you many many times more than a steamer. If you insist in using the MWO at least you will have to calibrate it. I recommed you to read Journal of Histotechnology 25(1):39-43 (2002). Rene J. Jeffrey Thompson wrote: Dear Histonetters, Can anyone recommend a good microwave for antigen retrieval? The cheap oven we have gives inconsistent temperatures that I don't want to rely on any longer. I haven't been able to find an adjustable wattage model that is not extremely high end, but I don't know if the adjustability is desirable but not absolutely essential. Any advice would be appreciated. I apologize if this is a hackneyed question and I haven't kept up. Thanks, Jeff Thompson _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From sjchtascp <@t> yahoo.com Mon Oct 10 11:51:18 2005 From: sjchtascp <@t> yahoo.com (Steven Coakley) Date: Mon Oct 10 11:51:39 2005 Subject: [Histonet] Brdu IHC Paraffin Message-ID: <20051010165118.28258.qmail@web90209.mail.scd.yahoo.com> Good afternoon, I need some "wise" insight into problems were having with our Brdu staining on paraffin embedded BDL rat liver. Instead of just the target cells staining around the bile ducts we got specific staining of all nuclei. Cytoplasam was clear. Any ideas. Steve --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From ploykasek <@t> phenopath.com Mon Oct 10 12:23:12 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Mon Oct 10 12:23:51 2005 Subject: [Histonet] microwave antigen retrieval In-Reply-To: Message-ID: Dear Jeff, Just my view, but to me the most important aspect of HIER is to standardize it as much as possible. If you are performing HIER in open coplin jars/dishes in the microwave, the best way to do this is to always use the same volume of liquid, the same # of slides, to do the ?hot spot?check of you microwave as discussed in various journal articles on the use of microwaves in the lab. That being said, I recommend using a steamer when possible and if you must microwave do so using a pressure cooker. For the steamer, you can standardize the size of container, the volume of liquid, and we insert thermometers through the holes in the lid of the steamer to standardize the temperature point that we start timing the HIER. For the microwave pressure cooker, we standardize the volume of liquid, the number of slides ran, and the time under pressure. The pressure cooker reaches pressure when the pressure indicator becomes raised, and there is an accompanying ?whistling? sound ? I will say that this is perhaps not true standardization as one has to become familiar with the correct ?whistle? sound, but it works for us. Well, I?ve rambled on about this enough , and haven?t even answered your question. I can tell you that we use an inexpensive microwave with the microwave pressure cooker. The only use for the microwave is for HIER using the pressure cooker. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA > Dear Histonetters, > > Can anyone recommend a good microwave for antigen retrieval? The cheap oven we > have gives inconsistent temperatures that I don't want to rely on any longer. > I haven't been able to find an adjustable wattage model that is not extremely > high end, but I don't know if the adjustability is desirable but not > absolutely essential. Any advice would be appreciated. I apologize if this is > a hackneyed question and I haven't kept up. Thanks, > > Jeff Thompson > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail message, including any attachments, is for the sole use of the intended recipients and may contain privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message, or you may call PhenoPath Laboratories, Seattle, WA U.S.A. at (206) 374-9000. From shahin.zangenehpour <@t> mail.mcgill.ca Mon Oct 10 12:33:42 2005 From: shahin.zangenehpour <@t> mail.mcgill.ca (Shahin Zangenehpour) Date: Mon Oct 10 12:34:03 2005 Subject: [Histonet] Un-mounting mounted tissue sections Message-ID: <2C5DF026-3AA3-4D3C-ADAC-FF62E2757C3B@mail.mcgill.ca> Dear Histonet-ers, Does anyone know of a way to un-mount paraformaldehyde fixed brain sections previously mounted on gelatin-subbed glass slides? I have been trying to optimise the IHC conditions for a particular antibody and in the process have run out of my free-floating sections. However, I had mounted some sections on glass for other purposes that I would like to un-mount (if possible) in order to complete my study. Any help you can provide is much appreciated. Best, SZ ________________________________________________________________________ _______________________________ Shahin Zangenehpour, PhD Postdoctoral Fellow | Montreal Neurological Institute | Cognitive Neuroscience Unit 3801 University Street | Room 276 | Montr?al QC Canada H3A 2B4 | P. 514 398 1717 | F. 514 398 1338 From emry <@t> u.washington.edu Mon Oct 10 14:11:15 2005 From: emry <@t> u.washington.edu (Trisha Emry) Date: Mon Oct 10 14:10:30 2005 Subject: [Histonet] tooth structure Message-ID: Hi, I have been requested to process a "special" tooth. I need some way to mark the position of the tooth throughout sectioning. Is there something I can embed and section along with the tooth to mark the architecture for computer imaging? I am doing this in paraffin. Thanks, Trisha U of WA Seattle From anh2006 <@t> med.cornell.edu Mon Oct 10 15:27:45 2005 From: anh2006 <@t> med.cornell.edu (Andrea Hooper) Date: Mon Oct 10 15:28:02 2005 Subject: [Histonet] microwave antigen retrieval Message-ID: <153344f15a90.434a9681@med.cornell.edu> I use a $15 veggie steamer that my DAKO rep gave me years ago (although I have replaced it since then with a similar model). It works AMAZINGLY using the DAKO Target Retrieval Solution with nearly no background and GREAT signal. The only drawback is time for heating up and the 20 minute retrieval and 20 minute cool down for a total of 1-1.5 hours retrieving. I am not a big fan of microwave techniques for retrieval as I have found my background to be worse and my signal to be less clear and sometimes inconsistent. But I agree with the other poster that consistency is the key to success no matter how it's achieved (likely through consistent use of positive and negative controls within each experiment). ------------------------------------------------------------------------------------------------------ > From: Jeffrey Thompson ? > Sent: Monday, October 10, 2005 11:53 am > To: histonet@lists.utsouthwestern.edu? > Subject: [Histonet] microwave antigen retrieval > > > >Dear Histonetters, > >Can anyone recommend a good microwave for antigen retrieval? The cheap oven we have gives inconsistent >temperatures that I don't want to rely on any longer. I haven't been able to find an adjustable wattage model that >is not extremely high end, but I don't know if the adjustability is desirable but not absolutely essential. Any advice >would be appreciated. I apologize if this is a hackneyed question and I haven't kept up. Thanks, > >Jeff Thompson From rjbuesa <@t> yahoo.com Mon Oct 10 15:41:25 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Mon Oct 10 15:41:41 2005 Subject: [Histonet] microwave antigen retrieval In-Reply-To: <153344f15a90.434a9681@med.cornell.edu> Message-ID: <20051010204125.54188.qmail@web61216.mail.yahoo.com> And that is exactly my point about the steamer and the microwave. To accelerate the process you can heat up the HIER solution in the microwave oven and transfer it to the steamer once it reaches the boliling point (100 Celsius). The steamer will give to an oustanding consistency and the microwave oven will speed the start of the HIER step (and consequently the whole process). In order to use the microwave oven to its fullest you should calibrate it first. And forget about the "hot spots": you can determine the hot spot in a MWO with a bulb array, but once you introduce a water load in it, the hot spot distribution will change completely (this according with Kok and Boon: "Microwaves for the art of microscopy", and with some experiment I have done). Rene J. Andrea Hooper wrote: I use a $15 veggie steamer that my DAKO rep gave me years ago (although I have replaced it since then with a similar model). It works AMAZINGLY using the DAKO Target Retrieval Solution with nearly no background and GREAT signal. The only drawback is time for heating up and the 20 minute retrieval and 20 minute cool down for a total of 1-1.5 hours retrieving. I am not a big fan of microwave techniques for retrieval as I have found my background to be worse and my signal to be less clear and sometimes inconsistent. But I agree with the other poster that consistency is the key to success no matter how it's achieved (likely through consistent use of positive and negative controls within each experiment). ------------------------------------------------------------------------------------------------------ > From: Jeffrey Thompson > Sent: Monday, October 10, 2005 11:53 am > To: histonet@lists.utsouthwestern.edu > Subject: [Histonet] microwave antigen retrieval > > > >Dear Histonetters, > >Can anyone recommend a good microwave for antigen retrieval? The cheap oven we have gives inconsistent >temperatures that I don't want to rely on any longer. I haven't been able to find an adjustable wattage model that >is not extremely high end, but I don't know if the adjustability is desirable but not absolutely essential. Any advice >would be appreciated. I apologize if this is a hackneyed question and I haven't kept up. Thanks, > >Jeff Thompson _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From anh2006 <@t> med.cornell.edu Mon Oct 10 16:26:48 2005 From: anh2006 <@t> med.cornell.edu (Andrea Hooper) Date: Mon Oct 10 16:27:04 2005 Subject: [Histonet] microwave antigen retrieval Message-ID: <16385c8f1384.434aa458@med.cornell.edu> Good idea. I like the pre-heating option and will try that next time especially if in a fix. To save time now, I have been putting the steamer on first thing in the morning before I start dehydrating etc. That way, by the time I am ready for HIER the steamer is heated up above 95 deg C and I am ready to go. From clarissabush <@t> sbcglobal.net Mon Oct 10 17:32:59 2005 From: clarissabush <@t> sbcglobal.net (CM Bush) Date: Mon Oct 10 17:33:15 2005 Subject: [Histonet] Calretinin and 10% NBF Fixed Frozen vs. Processed and Paraffin Embedded Brain Tissue Message-ID: <20051010223259.65399.qmail@web80311.mail.yahoo.com> Dear Histonet, We are trying to stain human anterior cingulate brain with anti Calretinin. We have taken a sample of said tissue (fixed in 10% NBF), processed and paraffin embedded one portion of the tissue and cut the other portion on a freezing stage microtome (fixed frozen sample cryo protected with 30% sucrose). The sections for both varieties mounted on slides. We have been able to get Calretinin staining (at 1:3000) for the processed, embedded tissue but not for the fixed frozen version of the same tissue. For all tissue, antigen retrieval was done using citrate buffer pH6 and microwaving 12 min high and 6 min low, in water filled, sealed secondary container. Also, we used TBS buffer with 0.4% Triton X for all sections. Would anyone have any ideas as to what might be going on that the processed/embedded tissue will stain and the fixed frozen version of the same sample won?t or hasn?t? Thank you very much, in advance. CMB From Barry.R.Rittman <@t> uth.tmc.edu Mon Oct 10 21:53:10 2005 From: Barry.R.Rittman <@t> uth.tmc.edu (Rittman, Barry R) Date: Mon Oct 10 21:56:18 2005 Subject: [Histonet] tooth structure Message-ID: Trisha Not totally sure what you want. It would help greatly if you let us know what parameters you wish to measure concerning the tooth's architecture. Are you concerned about distortion of the tooth during sectioning and mounting? Thanks Barry ________________________________ From: histonet-bounces@lists.utsouthwestern.edu on behalf of Trisha Emry Sent: Mon 10/10/2005 2:11 PM To: histo Subject: [Histonet] tooth structure Hi, I have been requested to process a "special" tooth. I need some way to mark the position of the tooth throughout sectioning. Is there something I can embed and section along with the tooth to mark the architecture for computer imaging? I am doing this in paraffin. Thanks, Trisha U of WA Seattle _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From SARAH.REEVES <@t> ekht.nhs.uk Tue Oct 11 02:21:20 2005 From: SARAH.REEVES <@t> ekht.nhs.uk (SARAH REEVES) Date: Tue Oct 11 02:22:36 2005 Subject: [Histonet] Masson Trichrome Message-ID: Can anyone help? We have just received our EQA results for special stains and gained a 4/10 for the Massons Trichrome. Just wondering what other SOP s are in practice that score well. Sarah Reeves ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. Any views or opinions presented are solely those of the author and do not necessarily represent those of East Kent Hospitals NHS Trust. If you are not the intended recipient, be advised that you have received this email in error and that any use, dissemination, forwarding, printing or copying of this email is strictly prohibited. If you have received this email in error please notify the system manager at the following email address: root.postmaster@ekht.nhs.uk www.kentandmedway.nhs.uk This footnote also confirms that although this email message has been swept by MIMEsweeper for the presence of computer viruses, it is strongly recommended that you carry out your own virus scan of this message and any attachments. www.mimesweeper.com ********************************************************************** From maramydave <@t> aol.com Tue Oct 11 08:33:01 2005 From: maramydave <@t> aol.com (maramydave@aol.com) Date: Tue Oct 11 08:33:18 2005 Subject: [Histonet] water for immuno Message-ID: <8C79C7F6F7BB37B-164C-1EEFC@MBLK-M02.sysops.aol.com> How does anyone define "appropriate" water for immunostaining. We use de-ionized and moniter it for bacteria, conductivity ,silicates- but what really defines good water for immuno? From pruegg <@t> ihctech.net Tue Oct 11 08:34:36 2005 From: pruegg <@t> ihctech.net (pruegg@ihctech.net) Date: Tue Oct 11 08:35:04 2005 Subject: [Histonet] tooth structure In-Reply-To: Message-ID: <200510111334.j9BDYfg6043395@pro12.abac.com> If you are just trying to maintain anatomy position through out processing, embedding, sectioning and staining, I use a system with samples where I will forinstance embed the superior side down with the posterior end pointing towards the label end of the embedding cassette, when I sectioning I make sure to take the section off the block without flipping it over and pickup the section so that the posterior end is pointing towards the label end of the slide each time. It takes a little concentration and you have to know how the tissue came out of the body (I am actually there so that it get oriented properly) but this works for me. I hope this helps and is what you mean. There are tattoo dyes you can use also to mark tissue. Patsy -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Rittman, Barry R Sent: Monday, October 10, 2005 8:53 PM To: Trisha Emry; histo Subject: RE: [Histonet] tooth structure Trisha Not totally sure what you want. It would help greatly if you let us know what parameters you wish to measure concerning the tooth's architecture. Are you concerned about distortion of the tooth during sectioning and mounting? Thanks Barry ________________________________ From: histonet-bounces@lists.utsouthwestern.edu on behalf of Trisha Emry Sent: Mon 10/10/2005 2:11 PM To: histo Subject: [Histonet] tooth structure Hi, I have been requested to process a "special" tooth. I need some way to mark the position of the tooth throughout sectioning. Is there something I can embed and section along with the tooth to mark the architecture for computer imaging? I am doing this in paraffin. Thanks, Trisha U of WA Seattle _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From lu_ze <@t> sbcglobal.net Tue Oct 11 11:57:46 2005 From: lu_ze <@t> sbcglobal.net (Ze Lu) Date: Tue Oct 11 11:59:58 2005 Subject: [Histonet] Aorta and coronary vessel sections source Message-ID: <010901c5ce84$e76380b0$5b02a8c0@OPTIMUM2> Hi, histonet friends, In one of project, we need to stain the aorta and coronary. We need a few these paraffine sections from different species including human. Is there any source that we can obtain or buy these paraffine sections? Any help or clue will be appreciated. Thanks. Ze Lu, Ph.D, Optimum Therapeutics, LLC From Charles.Embrey <@t> carle.com Tue Oct 11 12:32:47 2005 From: Charles.Embrey <@t> carle.com (Charles.Embrey) Date: Tue Oct 11 12:33:01 2005 Subject: [Histonet] Looking for Joe Golden Message-ID: Sorry about this, but I am looking for an Airforce friend of mine from Keesler AFB Histology Dept.. Keesler Med Center is still closed from Katrinia and I can't find him. Any contact info would be great. Thanks histonet, Charles Embrey PA(ASCP) Histology MGR Carle Clinic Illinois From BRIAN.CHELACK <@t> usask.ca Tue Oct 11 12:44:08 2005 From: BRIAN.CHELACK <@t> usask.ca (Brian Chelack) Date: Tue Oct 11 12:45:13 2005 Subject: [Histonet] Re: Water appropriate for IHC Message-ID: <434BF9E8.4208A85D@sask.usask.ca> A good rule of thumb for our lab is to use pyrogen free water for any application where enzyme activity is critical. We have a good ultrapure water system in our glassware and media prep lab, but every once in a while we had unexplainable declines in staining intensity. The only explanation that we could come up with was that these systems do not remove all pyrogens from the water(these can be small < 10,000 mw non-charged molecules that can inhibit HRPO or Alk phos reactions. These molecules can vary seasonally in the feedstock water, and their concentration in your lab water will also be dependent on how well the water purification system is maintained. Since we have changed over to using pyrogen free water (used for surgical irrigation), we have had fewer of these types of unexplained staining isses (not to say that we don't have other staining issues though...) This type of water is not cheap, but neither are stain repeats. Brian Chelack Prairie Diagnostic Services From wlove <@t> att.net Tue Oct 11 13:52:29 2005 From: wlove <@t> att.net (wlove@att.net) Date: Tue Oct 11 13:52:47 2005 Subject: [Histonet] Microwave use Message-ID: <101120051852.22739.434C09ED0005B86C000058D321612436460A90010499@att.net> Hello, this is my first response to a Histonet discussion - what an excellent method to communicate - I do not believe I have seen anything like it. However, as I am unfamiliar with it, I am not sure that this will reach the Histonet. We do research and provide microwave heating and processing devices for laboratories and light industrial applications including research tissue processors and other microwaves for staining, etc... Microwaves are great time savers as the energy adsorbs into the material being heated differently that the way conventional heating does. They can be quite consistent as well once people understand how to use them and how they operate. Looking for hot spots with a bulb array is not a great method for finding hot spots as Rene J. pointed out. She accurately also pointed out that the hot spot will change with a load in it. By the way, if you can control power well, you do not need a separate water load. But the introduction of any material in the microwave cavity will certainly change the energy distribution pattern and thus the so called hot spot. Consistency is important to consistent processing. Factors that affect consistency include: Line voltage - always use a microwave oven that compensates for line voltage changes. Most do. Load - do not use a 50 ml sample and think you will get the same results with a 500 ml sample. Nor will 10 - 50 ml samples vs. a 500 ml sample as the samples will be in different locations within the cavity and thus change the energy density. Starting temperature of the sample will make differences in results Location in the microwave will change results. Mark with a marker a location and always use it. I have heard of other methods to get consistent results as I am sure you have. Here is one that I have never heard with regard to histoprocessing. The temperature of the magnetron and the power supply transformer (major microwave generator components) makes just about the biggest single differences (can be over 10% variations) in process consistency. The heating of the magnetron is quick, in about 3 minutes of operation, but the transformer can take 15 minutes or more as it is a large thermal mass. One way to reduce the variation is to operate the microwave with a liter or two of water on full power for 20-30 minutes to pre-warm the microwave. The other way is to use a microwave that uses the temperature of the sample to control the process. We are not biologist but are microwave engineers and understand the operation of microwaves and how they heat. Hope this makes sense to all of you. Caution - If I understand it correctly, you are using the microwave to heat the HIER solution to boiling and then putting it into the steamer. This would greatly decrease the initial heating time. Be very careful as it is possible to superheat, but not boil liquids in a microwave and then have it vigorously boil explosively when the liquid is move or stirred causing potential injuries. Tips to prevent: Do not use a container with a small mouth! Stir the solution every couple of minutes. Leave the solution in the container undisterbed for a period of time before removing. Protect you face and body (arms and hands also) with face shields nad proper clothing Best regards Wayne Love Microwave Research and Applications, Inc. 8685 Cherry Lane Laurel, MD 20707 Phone 301-953-1771 Fax 301-369-0523 WebSite www.microwaveresearch.com Email info@microwaveresearch.com Cell 630-269-5158 Direct email: wlove@att.net From jcox90 <@t> yahoo.com Tue Oct 11 14:23:42 2005 From: jcox90 <@t> yahoo.com (Jill Cox) Date: Tue Oct 11 14:24:09 2005 Subject: [Histonet] Cassette Labeler Message-ID: <20051011192343.43419.qmail@web52111.mail.yahoo.com> Hello Histonetters, We will be purchasing a cassette labeler soon and hopefully slide labeler down the road. I would love your feedback on which one's are more reliable. I have used a Shurmark in the past and it seemed to work fine. If there is something else out there you are using I would like to look into that as well. Thank you in advance, Jill Jill Cox HT (ASCP) From eckjm <@t> yahoo.com Tue Oct 11 22:38:36 2005 From: eckjm <@t> yahoo.com (john eckman) Date: Tue Oct 11 22:38:46 2005 Subject: [Histonet] 37% formalin or penfix and false negative ER/PR stains Message-ID: <20051012033836.64027.qmail@web60019.mail.yahoo.com> Can anyone comment on what effect using 37% formalin to fix breast tissue has on ER/PR immuno stains? Same question regarding alcoholic formalin use. We work with pathology residents and have never ending problems with under fixed breast tissue. Some have suggested using 37% formalin or an alcoholic formalin. We're concerned about these fixatives altering immuno stains, although some in our department have used 37% formalin elsewhere without stain issues. Our breast sections usually fix in 10% buffered formalin for 6 to 24 hours before processing and we still get underprocessed/underfixed blocks in histology. These blocks are then sent through the tissue processor again (new Themo processor). Could the re-processing alter our ER/PR stains as well? Appreciate any comments. John --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From christelle.gerard <@t> novartis.com Wed Oct 12 04:04:12 2005 From: christelle.gerard <@t> novartis.com (christelle.gerard@novartis.com) Date: Wed Oct 12 04:04:34 2005 Subject: [Histonet] anti Rat MMP2 or MMP14 Message-ID: Dear All, I m looking for primary antibody anti Rat MMP12 or MMP14, which works on parraffin tissue fixed 10% formalin. So if you can recommand me a vendor, a clone. Thanks you for you answer. Christelle From andromeda_tm <@t> libero.it Wed Oct 12 04:40:39 2005 From: andromeda_tm <@t> libero.it (Massimo) Date: Wed Oct 12 04:40:57 2005 Subject: [Histonet] amphibian histology atlas ... Message-ID: <001701c5cf11$027b5d00$beb01997@SN300208440005> Hi All, Does anybody know where I could find, on the Net, a free amphibian anatomy/histology atlas? Or some web site where could I collect information on amphibian and their embryo tissue histology? Thanks in advance. Massimo From Halynn <@t> aol.com Wed Oct 12 08:50:37 2005 From: Halynn <@t> aol.com (Halynn@aol.com) Date: Wed Oct 12 08:50:55 2005 Subject: [Histonet] Regulations in Kentucky Message-ID: <1c8.331594b6.307e6ead@aol.com> I'm doing a research project for one of our residents at the University of South Florida and am looking for the rules and regulation for that state. Unlike Florida's web site, the state of Kentucky does not list on their government page anything about medical quality assurance. If anyone on the list works in Kentucky and knows where to find the regulations pertaining to their state, I would sure appreciate the info. Cheryl Bodden University of South Florida From rjbuesa <@t> yahoo.com Wed Oct 12 09:48:35 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Wed Oct 12 09:48:52 2005 Subject: [Histonet] 37% formalin or penfix and false negative ER/PR stains In-Reply-To: <20051012033836.64027.qmail@web60019.mail.yahoo.com> Message-ID: <20051012144835.14417.qmail@web61222.mail.yahoo.com> 37% formalin (formaldehyde or methanal) is the "pure" substance. Being a GASS formalin combines with water up to a concentration of 37%; there is no stronger solution. The regular "10% formalin" is a dilution 1:10 of the 37% and, therefore, it is in reality a 3.7% solution of the natural gass methanal. Having said that, the difficulty with fixation with formalin on IHC resides in the fact that formalin fixes by cross-linking proteins (this is the nature of the epitopes) and HIER (Heat Induced Epitope Retrieval) is aimed at "un-crosslinking" at "undoing" the mess that formalin causes. Now, since formalin is an aldehyde, it can oxidize with the oxygen in the air, and when that happens it transforms into an acid (mathanoic or formic acid in this case), and this is why the "10%" formalin has to be neutralized with salts that will act as a buffer and maintain the pH of the "10%" formalin within "neutral" (non acid) levels. The problems you are having with breast tissue will NOT be solved by increasing the formalin concentration; by doing so you will create a stronger crosslinkage that will not be reversed during a "normal" HIER and this could explain why ER/PR detection does not take place. The problem will be solved with neutral buffered "10%" formal (NBF) acting MORE time (the 24 hours you said are OK) and on THINNER breast sections. Also, if you want to be sure that your ER/PR protocol is working correctly, do not use breast as your positive control; use CERVIX which is better, more stable, and consistent. I would avoid alcoholic formalin (not necessary and of unknown effect on epitopes). Check you processing protocol; if the tissue is well fixed and the processing is incorrect you will end with poorly infiltrated tissue that will cause meny problems with the detection (specially if your sections are not used for IHC immediately). There is not such a thing as "reprocessing"; once the tissue is processed improperly it cannot be "resurrected". Hope this will help! Rene J. john eckman wrote: Can anyone comment on what effect using 37% formalin to fix breast tissue has on ER/PR immuno stains? Same question regarding alcoholic formalin use. We work with pathology residents and have never ending problems with under fixed breast tissue. Some have suggested using 37% formalin or an alcoholic formalin. We're concerned about these fixatives altering immuno stains, although some in our department have used 37% formalin elsewhere without stain issues. Our breast sections usually fix in 10% buffered formalin for 6 to 24 hours before processing and we still get underprocessed/underfixed blocks in histology. These blocks are then sent through the tissue processor again (new Themo processor). Could the re-processing alter our ER/PR stains as well? Appreciate any comments. John --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From Myri37 <@t> aol.com Wed Oct 12 10:56:17 2005 From: Myri37 <@t> aol.com (Myri37@aol.com) Date: Wed Oct 12 10:56:35 2005 Subject: [Histonet] remove grinding debris Message-ID: <0187B074.2F7C3685.0005167B@aol.com> Hello everyone, I grind on abrasive sandpaper, sections of titanium implants embedded in epon. Titanium is coated with a hydroxyapatite layer and soft tissue. After staining with RBS (rapid bone stain), i see on slides a lot of debris,i think they are grinding debris and titanium or hydroxyapatite dust... Does anyone have a process to remove these debris ? Do you recommend a wash with a specific detergent solution after grinding, does 0.1 % Zephiran chloride solution efficient ? Thank you very much for any advice. Myriam NI France From pruegg <@t> ihctech.net Wed Oct 12 11:03:36 2005 From: pruegg <@t> ihctech.net (Patsy Ruegg) Date: Wed Oct 12 11:03:56 2005 Subject: [Histonet] An old tissue processor Message-ID: <200510121603.j9CG3Trx019988@chip.viawest.net> We have an old Fisher Histomatic Model 166A tissue processor that is on the fritz. Does anyone know where we might be able to find parts and service for such a critter???? Best regards, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. From Ronnie_Houston <@t> bshsi.com Wed Oct 12 11:10:38 2005 From: Ronnie_Houston <@t> bshsi.com (Houston, Ronnie) Date: Wed Oct 12 11:11:00 2005 Subject: [Histonet] MOUSE BRAIN FIXATION Message-ID: <530361BF03351B4CAE5270A05D3037B506B71A9B@bsrexms01.bshsir.com> There have been several inquiries regarding fixation of mouse brains recently. A short technical report has just been published in BioTechniques: Minimally invasive method for murine brain fixation Eichenbaum KD et al Biotechniques 2005; 39(4): 487-500 After simple registration, it can be accessed at http://www.biotechniques.com Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com ________________________________________________________________________________________________________________________________ ________________________________________________________________________________________________________________________________ The information in this communication is intended to be confidential to the Individual(s) and/or Entity to whom it is addressed. It may contain information of a Privileged and/or Confidential nature, which is subject to Federal and/or State privacy regulations. In the event that you are not the intended recipient or the agent of the intended recipient, do not copy or use the information contained within this communication, or allow it to be read, copied or utilized in any manner, by any other person(s). Should this communication be received in error, please notify the sender immediately either by response e-mail or by phone, and permanently delete the original e-mail, attachment(s), and any copies. From judi.ford <@t> jax.org Wed Oct 12 11:44:03 2005 From: judi.ford <@t> jax.org (judi.ford@jax.org) Date: Wed Oct 12 11:45:34 2005 Subject: [Histonet] Maine Society for Histotechnology Meeting Message-ID: <8327654.1129135443554.JavaMail.ocsadmin@jcs-mid-prod.jax.org> Hi everyone, I just wanted to let everyone know that our society here in Maine will be holding their annual meeting for this year here at the Jackson Laboratory in Bar Harbor on Saturday, October 22. We have three speakers giving talks on subjects such as "Staining techniques in Research", "Advances in Microscopy" and " Using the Dako Autoimmunostainer in Research". Fall is in full bloom up here and the stores in town are having their close out sales. Its a great time to visit! If you have any questions don't hesitate to give us a call (207)288-6193. Sorry for the late notice :( Judi Ford HIstotechnologist HIstopathology & Microscopy The Jackson Laboratory 600 Main St. Bar Harbor, Me 04609 207-288-6193 From tmhpath <@t> amigo.net Wed Oct 12 11:49:31 2005 From: tmhpath <@t> amigo.net (Michelle D. Moore) Date: Wed Oct 12 11:47:57 2005 Subject: [Histonet] Cytologies crossing state lines Message-ID: <000901c5cf4c$eb22d5c0$ef00a8c0@tmhpath1> Hello in histoland. I am looking for information on whether or not you can ship freshly collected pap smears across state lines?! I have been told you can by one place and then told that you cannot at all. I know that you cannot send them across California's state line but what about the rest of the nation? I appreciate any help I can get. Thank you for your time and help. Michelle D. Moore The Memorial Hospital Craig, CO tmhpath@amigo.net From ttroyer <@t> petersonlab.com Wed Oct 12 11:58:11 2005 From: ttroyer <@t> petersonlab.com (Travis Troyer) Date: Wed Oct 12 11:58:24 2005 Subject: [Histonet] Air Bubbles in Manual Coverslipping Message-ID: <003501c5cf4e$217bb000$6601010a@Peterson.local> We have recently come under the "gun" for excess bubbles and glue on the slides after coverslipping. We currently do all of coverslipping manually and was wondering if anyone had any helpful hints to keep the bubbles out and and excess glue off the slide. Thanks for the assistance, Travis Troyer Peterson Laboratory Services From PIXLEYSK <@t> UCMAIL.UC.EDU Wed Oct 12 12:20:52 2005 From: PIXLEYSK <@t> UCMAIL.UC.EDU (Pixley, Sarah (pixleysk)) Date: Wed Oct 12 12:21:01 2005 Subject: [Histonet] RE: use of microwaves Message-ID: Dear All: I wondered if anyone that is currently using microwaves to heat samples has considered using a sand dry-bath for heating? The molecular biologists have all switched to those and they are really great. They are just heaters with a well that you put a good quality sand in. They can heat the sand up to 130 degrees centigrade. Then you stick your sample in the sand. The heat is very consistent. You can have something go to close to boiling almost instantly, without the muss and fuss of a hot water bath (or a microwave). Aside from a few bits of sand getting out, they are very civilized and clean. And they take up FAR less counter space than a microwave. Just a thought, but it is from someone who is not doing HIER. Sarah From rjbuesa <@t> yahoo.com Wed Oct 12 12:22:58 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Wed Oct 12 12:23:12 2005 Subject: [Histonet] Air Bubbles in Manual Coverslipping In-Reply-To: <003501c5cf4e$217bb000$6601010a@Peterson.local> Message-ID: <20051012172258.72021.qmail@web61225.mail.yahoo.com> You should start with the fluidity of the "glue" (I always prefer to call it "mounting medium"). It should be fluid enough to spread over the section. You should apply a very small drop over the section with either a plastic dropper or a wood applicator. The coverslip should be over a paper towel and you should get the slide/section with the mounting medium drop on it in contact with the coverslip and press it gently. You should wrap your finger with a cotton fiber disposable gauze to immediately clean the excess mounting medium. If the mounting medium is liquid enough you will prevent the bubbles, and even if the amount is in excess you can clean it around the coverslip eliminating the "overflow". Rest the coverslipped slide flat to dry out. Dammar resin or any other commercial mounting medium will do. Regardless if the diluent is toluene or xylene, you can always adjust the fluidity with xylene (which is totally miscible with toluene). Practice is essential ("like getting to Carnegie Hall!"). Hope this will help. Rene J. Travis Troyer wrote: We have recently come under the "gun" for excess bubbles and glue on the slides after coverslipping. We currently do all of coverslipping manually and was wondering if anyone had any helpful hints to keep the bubbles out and and excess glue off the slide. Thanks for the assistance, Travis Troyer Peterson Laboratory Services _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From Barry.R.Rittman <@t> uth.tmc.edu Wed Oct 12 13:18:31 2005 From: Barry.R.Rittman <@t> uth.tmc.edu (Rittman, Barry R) Date: Wed Oct 12 13:18:40 2005 Subject: [Histonet] remove grinding debris Message-ID: Myriam. Are you using water as a lubricant on the paper? If so remember that Epon is somewhat hygroscopic. Can grind using a neutral oil. If grinding dry then repeat the grinding on a frosted glass plate. This often will remove a lot of the debris. Can first try a commercial frosted slide or prepare your own using alumina. Are you sure that the debris is not being introduced during staining? Hydroxyapatite and other minerals can cause precipitates and also may bind to some dyes. Barry -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Myri37@aol.com Sent: Wednesday, October 12, 2005 10:56 AM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] remove grinding debris Hello everyone, I grind on abrasive sandpaper, sections of titanium implants embedded in epon. Titanium is coated with a hydroxyapatite layer and soft tissue. After staining with RBS (rapid bone stain), i see on slides a lot of debris,i think they are grinding debris and titanium or hydroxyapatite dust... Does anyone have a process to remove these debris ? Do you recommend a wash with a specific detergent solution after grinding, does 0.1 % Zephiran chloride solution efficient ? Thank you very much for any advice. Myriam NI France _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From sharon.osborn <@t> dnax.org Wed Oct 12 13:49:25 2005 From: sharon.osborn <@t> dnax.org (Osborn, Sharon) Date: Wed Oct 12 13:50:12 2005 Subject: [Histonet] slide, cassette labelers Message-ID: <29B25753F6B1D51196110002A589D44402398230@PALMSG30.us.schp.com> Jill, Please consider the Leica cassette and slide printers (not labelers). We have had ours almost 1 year with multiple users. They are very good, consistent and the service has been great. They are just about ready for a major upgrade to solve some problems that developed this past year with units out in the field. Still with those problems, these are better than the previous ones. As you, I have used the ShurMark for years...it was great for its time and definitely filled a void. However, the Leica is the new generation of labelers and well worth your having it demonstrated in your facility. Sharon Osborn DNAX, Schering Plough BioPharma Palo Alto, CA 650.496.6539 Date: Tue, 11 Oct 2005 12:23:42 -0700 (PDT) From: Jill Cox Subject: [Histonet] Cassette Labeler To: histonet@lists.utsouthwestern.edu Message-ID: <20051011192343.43419.qmail@web52111.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Hello Histonetters, We will be purchasing a cassette labeler soon and hopefully slide labeler down the road. I would love your feedback on which one's are more reliable. I have used a Shurmark in the past and it seemed to work fine. If there is something else out there you are using I would like to look into that as well. Thank you in advance, Jill Jill Cox HT (ASCP) ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. From JSCHUMA1 <@t> Fairview.org Wed Oct 12 14:02:22 2005 From: JSCHUMA1 <@t> Fairview.org (Schumacher, Jennifer J) Date: Wed Oct 12 14:02:39 2005 Subject: [Histonet] slide, cassette labelers Message-ID: I agree completely. Our slide labeler was down one day, and we tried to use our old ShurMark as a backup. Only then did we remember how lucky we were to have the new units. They are cleaner, easier to use, and oh so FAST!!! Plus, we are saving a fortune by not buying the "colormark" slides any longer. I would highly recommend them. Jennifer Schumacher, University of Minnesota Medical Center, Fairview -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Osborn, Sharon Sent: Wednesday, October 12, 2005 1:49 PM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] slide, cassette labelers Jill, Please consider the Leica cassette and slide printers (not labelers). We have had ours almost 1 year with multiple users. They are very good, consistent and the service has been great. They are just about ready for a major upgrade to solve some problems that developed this past year with units out in the field. Still with those problems, these are better than the previous ones. As you, I have used the ShurMark for years...it was great for its time and definitely filled a void. However, the Leica is the new generation of labelers and well worth your having it demonstrated in your facility. Sharon Osborn DNAX, Schering Plough BioPharma Palo Alto, CA 650.496.6539 Date: Tue, 11 Oct 2005 12:23:42 -0700 (PDT) From: Jill Cox Subject: [Histonet] Cassette Labeler To: histonet@lists.utsouthwestern.edu Message-ID: <20051011192343.43419.qmail@web52111.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Hello Histonetters, We will be purchasing a cassette labeler soon and hopefully slide labeler down the road. I would love your feedback on which one's are more reliable. I have used a Shurmark in the past and it seemed to work fine. If there is something else out there you are using I would like to look into that as well. Thank you in advance, Jill Jill Cox HT (ASCP) ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rchiovetti <@t> aol.com Wed Oct 12 14:03:44 2005 From: rchiovetti <@t> aol.com (rchiovetti@aol.com) Date: Wed Oct 12 14:04:12 2005 Subject: [Histonet] An old tissue processor In-Reply-To: <200510121603.j9CG3Trx019988@chip.viawest.net> References: <200510121603.j9CG3Trx019988@chip.viawest.net> Message-ID: <8C79D76CD48E79F-17C4-3A40@MBLK-M07.sysops.aol.com> Patsy, Boy that *is* an oldie! The Fisher products were taken over by RMC, Inc. several years ago. Then RMC sold the product line to Ventana Medical Systems here in Tucson. It's a long shot, but you might try calling Ventana at (800) 227-2155. Ask for Tom Kennedy. Tom's was with the product line from the original Fisher days, then RMC, then Ventana. He would know if anyone would about parts for the 166. Good Luck! Cheers, Bob Chiovetti The Microscope works Arizona's Microscopy Resource Tucson, Arizona USA Tel./Fax 520-546-4986 Member, Arizona Small Business Association (www.asba.com) -----Original Message----- From: Patsy Ruegg To: histonet@pathology.swmed.edu Sent: Wed, 12 Oct 2005 10:03:36 -0600 Subject: [Histonet] An old tissue processor We have an old Fisher Histomatic Model 166A tissue processor that is on the fritz. Does anyone know where we might be able to find parts and service for such a critter???? Best regards, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From sharon.osborn <@t> dnax.org Wed Oct 12 14:03:52 2005 From: sharon.osborn <@t> dnax.org (Osborn, Sharon) Date: Wed Oct 12 14:05:16 2005 Subject: [Histonet] Coverslipping technics Message-ID: <29B25753F6B1D51196110002A589D44402398231@PALMSG30.us.schp.com> Travis, One of my favorite tasks is coverslipping. However, due to pressing duties and volume, the automated coverslipper does the work. To get a good coverslip technic is essential when doing so manually. This is the one I use: 1. The slide is moist from the xylene. I use a dropper (I do have one of the old type bottles with the little glass dropper with a round glass bulb on its end)or a wooden applicator stick. Dip the stick into xylene to moisten it then into the mounting medium. The mounting medium will form a ball drop on the applicator. 2. Place the ball drop of mounting medium directly on the lower edge of the slide. I generally make a little line of it along the edge. 3. Pick up the coverslip with two fingers(I use thumb and index) along the coverslip edges. Be certain there are no sticky coverslips stuck together. Place the long edge of the coverslip along the lower edge of the slide on the mounting medium. The slide will gently lay down on the mounting medium and move it over the slide as it makes contact. There are rarely any bubbles. 4. If there are bubbles, gentle pressure with forceps or the applicator end (not one with mounting medium) will move those out. Also, there is rarely any excell medium around the edges. I 5. If there is excess mountimg medium around the edges, wipe it gently with a Kimwipe moistened with xylene or use a #3 artist brush (camel hair) dipped in xylene to clean the edges of the slide. Lay in mats to dry. Practise will develop this technic such that you will be faster than the automated coverslipper--the glass ones--which are all I recommend!..:-) Sharon Osborn DNAX, Schering Plough BioPharma Palo Alto, CA 650.496.6539 Date: Wed, 12 Oct 2005 11:58:11 -0500 From: "Travis Troyer" Subject: [Histonet] Air Bubbles in Manual Coverslipping To: Message-ID: <003501c5cf4e$217bb000$6601010a@Peterson.local> Content-Type: text/plain; charset="iso-8859-1" We have recently come under the "gun" for excess bubbles and glue on the slides after coverslipping. We currently do all of coverslipping manually and was wondering if anyone had any helpful hints to keep the bubbles out and and excess glue off the slide. Thanks for the assistance, Travis Troyer Peterson Laboratory Services ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. From rjbuesa <@t> yahoo.com Wed Oct 12 15:45:52 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Wed Oct 12 15:46:11 2005 Subject: [Histonet] RE: use of microwaves In-Reply-To: Message-ID: <20051012204552.15123.qmail@web61225.mail.yahoo.com> I have used sand bath during some biochemistry procedures and I agree with you that they are great BUT (there is always a BUT) temperature regulation is somewhat tricky and will depend on the amount of sand and the condition of the heating elements. For HIER you will have to consider the following aspects: 1- it requires almost one half an hour to complete. 2- How much HIER solution are you going to use? If not enough it will probably evaporate while in the sand bath after just a few minutes. 3- evaporation is prevented in the steamer because the environment is water vapor saturated and the vapor phase is, lets say "compensated". 4- in the microwave oven you will limit evaporation because HIER is quicker (due to the effect of the microwaves on the bipolar molecules within the tissue, and this, along with heat, is the fundament for the microwave oven). You will have to do many tests to find out an ideal ratio volume/temperatureXtime for a HIER done in a sand bath. The steamer is so simple to operate, and so consistent, that I personally would not try the sand bath to do HIER. This is just my opinion. Hope this will help! Rene J. "Pixley, Sarah (pixleysk)" wrote: Dear All: I wondered if anyone that is currently using microwaves to heat samples has considered using a sand dry-bath for heating? The molecular biologists have all switched to those and they are really great. They are just heaters with a well that you put a good quality sand in. They can heat the sand up to 130 degrees centigrade. Then you stick your sample in the sand. The heat is very consistent. You can have something go to close to boiling almost instantly, without the muss and fuss of a hot water bath (or a microwave). Aside from a few bits of sand getting out, they are very civilized and clean. And they take up FAR less counter space than a microwave. Just a thought, but it is from someone who is not doing HIER. Sarah _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Wed Oct 12 15:49:14 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Wed Oct 12 15:49:26 2005 Subject: [Histonet] Cytologies crossing state lines In-Reply-To: <000901c5cf4c$eb22d5c0$ef00a8c0@tmhpath1> Message-ID: <20051012204914.21514.qmail@web61212.mail.yahoo.com> I would contact UPS or FederalExpress. Some time ago we had to send some samples and they knew all the regulations/limitations (since they are the carriers). Rene J. "Michelle D. Moore" wrote: Hello in histoland. I am looking for information on whether or not you can ship freshly collected pap smears across state lines?! I have been told you can by one place and then told that you cannot at all. I know that you cannot send them across California's state line but what about the rest of the nation? I appreciate any help I can get. Thank you for your time and help. Michelle D. Moore The Memorial Hospital Craig, CO tmhpath@amigo.net _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From cbass <@t> bidmc.harvard.edu Wed Oct 12 16:19:07 2005 From: cbass <@t> bidmc.harvard.edu (Caroline Bass) Date: Wed Oct 12 16:19:24 2005 Subject: [Histonet] mounting media for EGFP in liver Message-ID: <41489ADD-7323-463D-88F8-42BBAC1B341A@bidmc.harvard.edu> Hello, I have used a viral vector to transduce liver and pancreatic tissues in the mouse with EGFP and RFP. I have cut portions of the liver and pancreas into small blocks and fixed overnight with NBF. What is the best way to visualize the signal? I want to cut 30 micron sections, as I only have access to a sliding microtome. Could someone recommend a good, and hopefully inexpensive, mounting media for fluorescence? I have a fairly strong signal but I am worried about losing it if I use the wrong product. I am not very experienced with fluorescence so I really don't know what to use. Someone recommended Aqua PolyMount from polysciences. Any suggestions would be appreciated. Caroline From froyer <@t> bitstream.net Wed Oct 12 16:19:09 2005 From: froyer <@t> bitstream.net (Ford Royer) Date: Wed Oct 12 16:19:33 2005 Subject: [Histonet] Cytologies crossing state lines In-Reply-To: <000901c5cf4c$eb22d5c0$ef00a8c0@tmhpath1> Message-ID: I believe that it is okay, as long as it is not for an immoral purpose... ...sorry, couldn't resist. ;-) (For you youngsters out there... see "The Mann Act of 1910") ~ Ford Ford Royer, MT(ASCP) Minnesota Medical Specialists 7177 Madison Ave. W. Golden Valley, MN 55427-3601 888-790-9686 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Michelle D. Moore Sent: Wednesday, October 12, 2005 11:50 AM To: Histonet Subject: [Histonet] Cytologies crossing state lines Hello in histoland. I am looking for information on whether or not you can ship freshly collected pap smears across state lines?! I have been told you can by one place and then told that you cannot at all. I know that you cannot send them across California's state line but what about the rest of the nation? I appreciate any help I can get. Thank you for your time and help. Michelle D. Moore The Memorial Hospital Craig, CO tmhpath@amigo.net _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Malcolm.McCallum <@t> tamut.edu Wed Oct 12 23:01:16 2005 From: Malcolm.McCallum <@t> tamut.edu (Malcolm McCallum) Date: Wed Oct 12 23:01:27 2005 Subject: [Histonet] decalcifying solutions for skeletochronology Message-ID: Hi, I am having some students do various skeletochronology projects with amphibians and reptiles. I could just buy some decalcifying solution, but we have so many acids, I figured I would make some. Any good recipes out there for decalcifying bone w/o heating? Malcolm L. McCallum Assistant Professor Department of Biological Sciences Texas A&M University Texarkana 2600 Robison Rd. Texarkana, TX 75501 O: 1-903-233-3134 H: 1-903-791-3843 Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html From jkiernan <@t> uwo.ca Wed Oct 12 23:57:47 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Wed Oct 12 23:58:08 2005 Subject: [Histonet] decalcifying solutions for skeletochronology References: Message-ID: <434DE94B.61DC3E1C@uwo.ca> Dear Dr McCallum, Every book in the field of histotechnology contains several recipes for decalcifying. The calcified material can be removed by dissolving in a suitable acid (usually hydrochloric or formic) or by chelation with a suitable anion (usually citrate or EDTA). The choice of method is based on the requirements of the investigation. If you are in doubt about choosing a method, ask the internet for references rather than recipes. There are scores of books and hundreds of papers! A solid place to start searxhing the literature is Pearse's Histochemistry. John A. Kiernan MB, ChB, PhD, DSc Professor, Dept of Anatomy & Cell Biology The University of Western Ontario London, Canada. ---------------------------- Malcolm McCallum wrote: > > Hi, > I am having some students do various skeletochronology projects with amphibians and reptiles. I could just buy some decalcifying solution, but we have so many acids, I figured I would make some. Any good recipes out there for decalcifying bone w/o heating? > > Malcolm L. McCallum > Assistant Professor > Department of Biological Sciences > Texas A&M University Texarkana > 2600 Robison Rd. > Texarkana, TX 75501 > O: 1-903-233-3134 > H: 1-903-791-3843 > Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html > From Eric <@t> ategra.com Wed Oct 12 19:58:54 2005 From: Eric <@t> ategra.com (Eric Dye (ext 223)) Date: Thu Oct 13 01:05:53 2005 Subject: [Histonet] Histotech Opportunites in your Area Message-ID: Histonetters I'm presently on a search for my best client companies seeking HistoTech Supervisors and Histo Bench Techs. These are mostly fulltime permanent positions. (I have a few travel/temp HistoTech positions as well - see below). Most of my Histo Techs positions are clinical, although I do have a few Research Histo Tech positions as well. Here are some of my Hottest jobs: 1. Louisiana( North Western) Openings for a HistoTech(Bench) No weekends, No call, and Top Dollar. 2. Ohio(Dayton Area) HistoTech Manager No Weekends, No call, Top Dollar if you are interested, please call me today at 1-800-466-9919 ext 223 3. Colorado (Greater Denver area) Opening for Several Bench Positions(Night Shift) No weekends, No call, Top Dollar 4. New Hampshire Openings for several Histotechs(Bench) No Weekends, No Call, Top Dollar, Excellent Benefits if you are interested, please call me today at 1-800-466-9919 ext 223 5. Minnesota Opening for HistoTech(Bench) No weekends, No Call, Top Dollar 6. New Jersey(Southern) Openings for HistoTech(Bench) No Weekends, No Call, Top Dollar if you are interested, please call me today at 1-800-466-9919 ext 223 7.Rhode Island Openings for Histotech(Bench) No weekends, No call, Top Dollar 8. California( Southern) HistoTech Supervisor(2nd shift) Openings for Histotechs(Bench,2nd shift) Great Location, No weekends, No call, Top Dollar 9. California(Southern) Openings for HistoTechs(Bench) Great Location, No weekends, No call, Top Dollar 10. Pennsylvania(Multiple jobs in greater Pittsburgh area) HistoTech opening(Bench) if you are interested, please call me today at 1-800-466-9919 ext 223 11. Illinois(Multiple jobs in Greater Illinois area) Opening for Histotech(Bench) 12. Michigan(Multiple jobs in Greater Michigan area) Openings for Histotechs(Bench) 13. Indiana Opening for Histotech(Bench) 14. Washington State(Eastern) Opening for Lead HistoTech(Bench) No weekends, No call if you are interested, please call me today at 1-800-466-9919 ext 223 15. Oklahoma Opening for Histotech(Bench) No weekends, No call. 16. Massachusetts (Boston Area) Part time Histo Tech(Permanent) No weekends, No call 17. Tennesee(Memphis area) Openings for Supervisor and Bench Techs 18. New Mexico openings for Supervisor and Bench Techs No Weekends, No Call, Excellent Benefits.. 19. Nebraska Openings for Histo Techs(Bench) No Weekends, No Call, Top Dollar 20. Virginia(Western Virginia) Opening for Bench Histotech No weekends, No call if you are interested, please call me today at 1-800-466-9919 ext 223 21. Ohio( Southern) Opening for Bench HistoTech No weekends, No call 22. Maryland( Baltimore area) Opening for Bench Histotech 23. Missouri(Greater Missouri area) Opening for Bench Histotech No weekends, No call 24. Wisconsin(Eastern area) Opening for a Bench Histotech No weekends, No call 25. Florida(Greater Florida area) Openings for Bench Histotech No weekends, No call The clients are currently interviewing candidates - so if you are interested, please call me today at 1-800-466-9919 ext 223 Thank you, Eric Dye-Sr Allied Healthcare Recruiter 1-800-466-9919 ext 223 From bmcmahill <@t> incytepathology.com Thu Oct 13 01:06:56 2005 From: bmcmahill <@t> incytepathology.com (Bonnie J. McMahill) Date: Thu Oct 13 01:07:20 2005 Subject: [Histonet] Height Adjustable tables Message-ID: Hi all, I was wondering if anyone has height adjustable (electronic or crank) tables for cutting? We are putting some in our lab, but are interested in purchasing some from a company that has a known track record. Thanks! Bonnie McMahill InCyte Pathology Spokane, WA From wilson_jm <@t> cimar.org Thu Oct 13 05:25:37 2005 From: wilson_jm <@t> cimar.org (Jonathan Wilson) Date: Thu Oct 13 05:15:51 2005 Subject: [Histonet] SGLT1 antibody References: <09E7875E806DFB4BBECBADBF966BDBB6F6763C@mksn79.d50.intra> <4346AA2E.FDDDFD38@uwo.ca> Message-ID: <003101c5cfe0$751b8950$17dea8c0@ciimar.up.pt> Hello, Could someone please recommend an antibody against SGLT1 (sodium glucose transporter) that works in rat or mouse FFPE tissue. I have tried a chemicon antibody but without any luck. Thanks for any advice. Sincerely, Jonathan Wilson CIIMAR-Ecofisiologia Rua dos Bragas 289 4050-123 Porto Portugal tel 351 22 340 1809 fax 351 22 339 0608 alt. e-mail: mop01258@mail.telepac.pt From ree3 <@t> leicester.ac.uk Thu Oct 13 06:27:10 2005 From: ree3 <@t> leicester.ac.uk (Edwards, R.E.) Date: Thu Oct 13 06:27:31 2005 Subject: [Histonet] RE: beta-2- microglobulin Message-ID: Looking for an antibody to the above that works, as ever, on paraffin processed mouse tissues... Many thanks Richard Edwards MRC TOX UNIT LEICESTER...U.K.... From rjbuesa <@t> yahoo.com Thu Oct 13 09:48:24 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 13 09:48:46 2005 Subject: [Histonet] decalcifying solutions for skeletochronology In-Reply-To: Message-ID: <20051013144824.30771.qmail@web61214.mail.yahoo.com> The decalcifying solution that you may end using (after any standard recipe you can find in any good histology manual) will depend on the type of bone you want to decalcify and what details you want to preserve. EDTA will give you the most details, but it will take longer; sulfuric acid will decalcify faster but will destroy almost any detail. The weaker the acid (lique citric) the slower the process but the more detail you will preserve; and an inverse correlation you will find with the stronger the acid. I hope this will help you to sort out your options. Rene J. Malcolm McCallum wrote: Hi, I am having some students do various skeletochronology projects with amphibians and reptiles. I could just buy some decalcifying solution, but we have so many acids, I figured I would make some. Any good recipes out there for decalcifying bone w/o heating? Malcolm L. McCallum Assistant Professor Department of Biological Sciences Texas A&M University Texarkana 2600 Robison Rd. Texarkana, TX 75501 O: 1-903-233-3134 H: 1-903-791-3843 Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From kaleid11 <@t> yahoo.com Thu Oct 13 10:19:46 2005 From: kaleid11 <@t> yahoo.com (Adam Perry) Date: Thu Oct 13 10:20:10 2005 Subject: [Histonet] MOUSE BRAIN FIXATION Message-ID: <20051013151946.35658.qmail@web30411.mail.mud.yahoo.com> I downloaded the technical report on this variant perfusion technique. I'm amazed that you can get good fixation of the brain with 1 ml of fixative pumped through the heart without clamping off descending aorta, etc. Has anyone tried this technique (or similar variants), and what kind of results were obtained? I'm planning on trying it in a week or so, because I would love to get good brain fixation with so little time, fixative and waste production...but am wary... Adam Perry Department of Physiology and Biophysics University of Illinois Chicago, IL 60612 There have been several inquiries regarding fixation of mouse brains recently. A short technical report has just been published in BioTechniques: Minimally invasive method for murine brain fixation Eichenbaum KD et al Biotechniques 2005; 39(4): 487-500 After simple registration, it can be accessed at http://www.biotechniques.com Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From cfavara <@t> niaid.nih.gov Thu Oct 13 10:35:32 2005 From: cfavara <@t> niaid.nih.gov (Favara, Cynthia (NIH/NIAID)) Date: Thu Oct 13 10:36:01 2005 Subject: [Histonet] MOUSE BRAIN FIXATION Message-ID: I have this paper and hope to try this next week. Note that the fixative is picric acid/paraformaldehyde/gluteraldehyde. I would love to get great fixation with 1ml and without opening the chest cavity. I will let you know! c Cynthia Favara NIAID/NIH/RML/LPVD 903 South 4th Street Hamilton, MT 59840 406-363-9317 Disclaimer: The information in this e-mail and any of its attachments is confidential and may contain sensitive information. It should not be used by anyone who is not the original intended recipient. If you have received this e-mail in error please inform the sender and delete it from your mailbox or any other storage devices. National Institute of Allergy and Infectious Diseases shall not accept liability for any statements made that are sender's own and not expressly made on behalf of the NIAID by one of its representatives -----Original Message----- From: Adam Perry [mailto:kaleid11@yahoo.com] Sent: Thursday, October 13, 2005 8:20 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] MOUSE BRAIN FIXATION I downloaded the technical report on this variant perfusion technique. I'm amazed that you can get good fixation of the brain with 1 ml of fixative pumped through the heart without clamping off descending aorta, etc. Has anyone tried this technique (or similar variants), and what kind of results were obtained? I'm planning on trying it in a week or so, because I would love to get good brain fixation with so little time, fixative and waste production...but am wary... Adam Perry Department of Physiology and Biophysics University of Illinois Chicago, IL 60612 There have been several inquiries regarding fixation of mouse brains recently. A short technical report has just been published in BioTechniques: Minimally invasive method for murine brain fixation Eichenbaum KD et al Biotechniques 2005; 39(4): 487-500 After simple registration, it can be accessed at http://www.biotechniques.com Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From amian <@t> thermage.com Thu Oct 13 11:32:24 2005 From: amian <@t> thermage.com (Arshia Mian) Date: Thu Oct 13 11:32:39 2005 Subject: [Histonet] Re: Tissue with Fat Message-ID: I have been having some problems with cutting Tissue with Fat on a cryostat (box temp and object temp are at -20C). The slices I cut tend to curl in the middle of the block close to the tissue. I have changed the blade several times, but it still continues to happen even with a new blade. Is there any correlation between the box temp and the object temp that would cause this to happen? Any suggestions? I have also noticed that at times, even though my setting is set to 20 microns, that I get thick and thin cuts. Any suggestions on this as well? I would greatly appreciate it. Thanks. From Charlotte.Kopczynski <@t> baycare.org Thu Oct 13 12:36:41 2005 From: Charlotte.Kopczynski <@t> baycare.org (Kopczynski, Charlotte) Date: Thu Oct 13 12:36:54 2005 Subject: [Histonet] Air Bubbles after Coverslipping Message-ID: <5452D66669CFC540B0452115ED26AF1F040E51E7@bcexch02.bcad.baycare.org> We have had problems at various times over the past 4-5 years. If you are manually coverslipping, make sure that your mounting medium is Xylene based if you are coverslipping from xylene. We had inadvertently tried to use a toluene based mounting medium. When we started using an automatic coverslipper, we had a huge problem. I found out that our coverglass was part of the problem. In order to keep coverglass from sticking together, it is covered with a very fine layer of ground glass. We found that bubbles were forming the next day after coverslipping. We converted to different coverglass and adjusted the amount of mounting media dispensed and no longer have the problem. I don't know if any of this will help but another thing to look at is how you are drying the slides after coverslipping. Are you having the bubble problem right after coverslipping or later after the slide has begun to dry? Thanks, Charlotte Kopczynski,HTL(ASCP) Baycare Pathology Manager Phone: 727-461-8246 Fax: 727-462-7597 Confidential: This electronic message and all contents contain information from BayCare Health System which may be privileged, confidential or otherwise protected from disclosure. The information is intended to be for the addressee only. If you are not the addressee, any disclosure, copy, distribution or use of the contents of this message is prohibited. If you have received this electronic message in error, please notify the sender and destroy the original message and all copies. From ccrowder <@t> vetmed.lsu.edu Thu Oct 6 10:28:08 2005 From: ccrowder <@t> vetmed.lsu.edu (Cheryl Crowder) Date: Thu Oct 13 13:18:58 2005 Subject: [Histonet] Bone Marrow Biopsies Message-ID: Hi all - I have a residen whose research will include doing bone marrow biopsies. Could someone help me with protocols for bone marrows. Vet Med here does not routinely do bone marrows and the results are spotty. What I am looking for is what you do at the point of biopsy, as make smears, fix some, etc., what fixative do you use, do you decal the BM and in what? All that "stuff". If anyone can help me, it would probably be better to e-mail me directly. Thank you, Cheryl Cheryl Crowder, BA, HTL(ASCP) Chief Technologist Anatomic Pathology Department of Pathobiological Sciences School of Veterinary Medicine Louisiana State University Skip Bertman Drive Baton Rouge, LA 70803 225-578-9734 FAX: 225-578-9720 From sgirees <@t> pathwaydx.com Thu Oct 6 11:36:50 2005 From: sgirees <@t> pathwaydx.com (Sherif Girees) Date: Thu Oct 13 13:18:59 2005 Subject: [Histonet] RE: Histonet Digest, Vol 23, Issue 7 Message-ID: <2BB61A9B5B12CF458B70EE8EF99A71A85B8EF4@PATHWAYMAIL1.PathwayCorp.local> To Patsy Ruegg, HT(ASCP)QIHC You have to anesthetize the animal first so the lungs will not claps, then open the chest cavity, make a small incision in the left and the ventricle, using a plastic syringe very gently inject PBS Ph 7.4 till the blood clears, the introduce the fix. We used that method to perfuse feline and swine as well as mice and rats for LT and EM. Sherif Girees,BS HT(ASCP)QIHC Manager Molecular Pathology Pathway Diagnostics 3003 Malibu Canyon Malibu CA.90265 Tel:310-774-3569 Fax:310-774-3556 s.girees@pathwaydx.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of histonet-request@lists.utsouthwestern.edu Sent: Thursday, October 06, 2005 7:59 AM To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 23, Issue 7 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit http://lists.utsouthwestern.edu/mailman/listinfo/histonet or, via email, send a message with subject or body 'help' to histonet-request@lists.utsouthwestern.edu You can reach the person managing the list at histonet-owner@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than "Re: Contents of Histonet digest..." Today's Topics: 1. Fetal mouse samples (Patsy Ruegg) 2. Re: Fetal mouse samples (Rene J Buesa) 3. 20 micron sections (cynthia haynes) 4. lymphatics (Hawkins, Hal K.) 5. Pax2 (Patti Loykasek) 6. RE: Positions in Louisiana (D. Ford) 7. RE: lymphatics (Luck, Greg D.) 8. Nevada Society of Histotechnology (connie grubaugh) 9. Re: lymphatics (John Kiernan) 10. Re: lymphatics (anh2006@med.cornell.edu) 11. Fixation using Modified Bouins and NBF (Ahmed, T (Tanni)) 12. RE: 20 micron sections (Lee & Peggy Wenk) 13. GFAP protocol for fluorescence (Stephen.Eyres@sanofi-aventis.com) 14. staining under hoods (Sauer, Barb) 15. RE: staining under hoods (Weems, Joyce) 16. amyloid in FNA smears (Houston, Ronnie) 17. High Complexity Testing - IHC (danaspears@frontiernet.net) 18. Metachromasia of mast cells in epoxy resin sections (gillian.2.brown@gsk.com) 19. Histology Journals (Prior, Andrew) 20. Re: 20 micron sections (Rene J Buesa) 21. Re: Histology Journals (Rene J Buesa) 22. Re: Nevada Society of Histotechnology (Rene J Buesa) 23. Job Openings In Dallas, Texas (Debbiejsiena@aol.com) 24. CAP question (Kapoor, Sue) 25. Region II Meeting November 4th and 5th, 2005 (Pamela Marcum) 26. AW: [Histonet] lymphatics (Gudrun Lang) ---------------------------------------------------------------------- Message: 1 Date: Wed, 5 Oct 2005 14:37:50 -0600 From: "Patsy Ruegg" Subject: [Histonet] Fetal mouse samples To: Message-ID: <200510052037.j95Kbf9O022163@chip.viawest.net> Content-Type: text/plain; charset="us-ascii" I am processing whole fetal mouse samples (these are pretty big, they have skin developed) and the investigator wants to have sections so that both lungs show up at the same time in the same section. Can someone recommend to me a way to dissect these to open them up so that they will fix and process well but still maintain the whole body architecture for sectioning. Thank you, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. ------------------------------ Message: 2 Date: Wed, 5 Oct 2005 14:00:18 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] Fetal mouse samples To: Patsy Ruegg , histonet@pathology.swmed.edu Message-ID: <20051005210019.5811.qmail@web61211.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Patsy: Many years ago I had to do that. Of the three "classical" sectioning planes (transverse, sagittal and frontal) the one to use is the frontal. Rest the embryo flat on its back and cut it open starting at nose/mouth level all the way back to the vertebral column in a way that the back half of the embryo can be separated from the front half. Try to harden it first with formaldehyde (4-6 hours will be enough; you should also inject in the abdomen a little amount of NBF also). Try to leave as much material in the front so you will get to cut to the lungs when trimming down the block. I recommend you also to process it enclosed in a cassette that keeps the structures in place, and to extend the times, not so much in the dehydrating as in the infiltrating steps. Had you been using the dehydrating/infiltrating steps I used to use (ethyl alcohol/isopropyl alcohol/mineral oil) you would get a better infiltration. Hope this will help. Rene J. Patsy Ruegg wrote: I am processing whole fetal mouse samples (these are pretty big, they have skin developed) and the investigator wants to have sections so that both lungs show up at the same time in the same section. Can someone recommend to me a way to dissect these to open them up so that they will fix and process well but still maintain the whole body architecture for sectioning. Thank you, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. ------------------------------ Message: 3 Date: Wed, 5 Oct 2005 14:14:29 -0700 (PDT) From: cynthia haynes Subject: [Histonet] 20 micron sections To: Histonet@lists.utsouthwestern.edu Message-ID: <20051005211429.83499.qmail@web33002.mail.mud.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Hello, everyone I am in need of information. I am cutting brain(human)tissue at 6 microns and 20 microns. The 6 microns sections are perfect. The 20 microns sections are an nightmare(and I mean Elm street nightmare). Can anyone suggest the best way to cut these sections without getting wrinkles and folds. I am at my wits end. Thanks in advance Cynthia Haynes H.T. ------------------------------ Message: 4 Date: Wed, 5 Oct 2005 16:15:48 -0500 From: "Hawkins, Hal K." Subject: [Histonet] lymphatics To: Message-ID: <8D6F233E2A5D574B929F3944F3316FD006310469@EXCH2K3.utmb.edu> Content-Type: text/plain; charset="us-ascii" For a model of necrotizing enterocolitis in the mouse we need to distinguish small veins from lymphatics in the gut. Could someone remind me of any immunostains or lectin that will allow us to tell the two apart? Many thanks, Hal Hawkins, UTMB Galveston ------------------------------ Message: 5 Date: Wed, 05 Oct 2005 14:53:42 -0700 From: Patti Loykasek Subject: [Histonet] Pax2 To: histonet Message-ID: Content-Type: text/plain; charset="ISO-8859-1" Hi all. I have a question about an antibody to Pax2. We have just begun to try to work up this antibody. It is supposedly upregulated in renal cell carcinoma, so it should be positive & it has been. From the reading I have done it should be negative in normal adult kidney. Our current problem is that we are seeing staining on normal adult kidney. Any help or ideas are appreciated. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information.? Any unauthorized review, use, disclosure or distribution is prohibited.? If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message or you may call PhenoPath Laboratories, Seattle, WA U.S.A at (206) 374-9000. ------------------------------ Message: 6 Date: Wed, 5 Oct 2005 17:21:44 -0500 From: "D. Ford" Subject: [Histonet] RE: Positions in Louisiana To: Message-ID: Content-Type: text/plain; charset="iso-8859-1" Delta Pathology, in Shreveport Louisiana, currently looking for HT/HTLs. If you have been displaced by the hurricanes and want to return to your home state or if you are looking to relocate please contact us for more information. Darlene Ford, B.S. CT, HT (ASCP) Technical Supervisor 2915 Missouri Avenue Shreveport, LA 71109 318-621-8820 phone 318-671-5922 fax dford@deltapathology.com ------------------------------ Message: 7 Date: Wed, 5 Oct 2005 15:44:14 -0700 From: "Luck, Greg D." Subject: RE: [Histonet] lymphatics To: "'Hawkins, Hal K.'" , Histonet@lists.utsouthwestern.edu Message-ID: Content-Type: text/plain Hal, Why don't you try the tools offered on www.immunoquery.com (this one is the best but you need to get a username and password) or www.IHCWorld.com. These are both excellent IHC resources. Greg Luck, BS, HT(ASCP) Anatomic Pathology Supervisor Deaconess Medical Center 800 W. 5th Ave Spokane, WA 99204 Phone 509.473.7077 Fax 509.473.7133 luckg@empirehealth.org www.deaconessmedicalcenter.org -----Original Message----- From: Hawkins, Hal K. [mailto:hhawkins@UTMB.EDU] Sent: Wednesday, October 05, 2005 2:16 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] lymphatics For a model of necrotizing enterocolitis in the mouse we need to distinguish small veins from lymphatics in the gut. Could someone remind me of any immunostains or lectin that will allow us to tell the two apart? Many thanks, Hal Hawkins, UTMB Galveston _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 8 Date: Thu, 06 Oct 2005 03:57:47 +0000 From: "connie grubaugh" Subject: [Histonet] Nevada Society of Histotechnology To: histonet@lists.utsouthwestern.edu Message-ID: Content-Type: text/plain; format="flowed" The Nevada Society of HIstotechnology will be having their next seminar Oct. 28th and 29th at Sunrise Hospital Auditorium in Las Vegas Nevada. There will be a Friday evening get together with the vendors. Drinks and snacks will be provided. This will start at 5:30 pm. Saturday sign in at 7 am. Coffee and rolls will be provided by Fisher Scientific. Meeting starts at 8 am. First meeting will be Pam Marcum -Is Your Tissue Processor Fighting You? Fight Back. This will be 3 ceu"s. There will be a luncheon provided by Sakura and Cardinal Health. A business meeting will also be held at this time. Afternoon meetings. 1 pm- Sandra L. Cummings, Putting the bug in Histotechnology, A basic understanding of Microorganisms. To follow Marianne Mailhoit- Exposure control plan for Blood Borne Pathogens. There will be no charge for this meeting. Hope to see everyone there. Vegas is the best place to be on Halloween weekend. Please email me with any questions, or call 702-396-4079 or 702-938-3619. Connie Grubaugh Nevada Society of Histotechnology Striving for Excellence Despite the Odds. ------------------------------ Message: 9 Date: Thu, 06 Oct 2005 00:18:11 -0400 From: John Kiernan Subject: Re: [Histonet] lymphatics To: "Hawkins, Hal K." Cc: Histonet@lists.utsouthwestern.edu Message-ID: <4344A583.842FBFA@uwo.ca> Content-Type: text/plain; charset=us-ascii Dear Hal Hawkins, The endothelium of lymphatic vessels can be stained by enzyme activity histochemistry. The enzyme is a 5-nucleotidase (belongs to the phosphatase family). Ordinarily this type of method is carried out with frozen sections of tissue that has been minimally fixed in formaldehyde or glutaraldehyde, or with unfixed cryostat sections that have been briefly fixed in cold acetone. The enzyme in lymphatics may be fairly robust because its activity can survive embedding in glycol methacrylate. The references below may help. If you choose one of these methods it will be important to consult the original publication because there's more than one sort of 5-nucleotidase. Each ref is followed by brief notes that I took when reading the papers. [An R in the acquisition number means I've got a Reprint of the whole paper; an A usually means I have a copy of the paper's Abstract. No letter with the acquisition number usually means I've seen the whole paper and took notes in the library.] 8194R. Kato,S; Yasunaga,A; Uchida,U (1991): Enzyme-histochemical method for identification of lymphatic capillaries. Lymphology 24, 125-129. Glycol methacrylate-embedded sections stained for 5'-nucleotidase & alkaline phosphatase. 5-N-ase in lymph capills; AlkP-ase in blood capills. 9239R. Nishida,S; Ohkuma,M (1993): Enzyme-histochemical staining of dermal lymphatic capillaries by guanylate cyclase. Lymphology 26, 195-199. Enzyme histochemical staining distinguishes lymph from blood capillaries. Says 5-nucleotidase & adenylate cyclase do so too. (Used unfixed cryostat sections.) 9665R. Okada,E (1994): An improved enzyme-histochemical method for identification of lymphatic capillaries on paraffin sections. Lymphology 27, Suppl, 732-735. Staining of blood & lymph capillaries. Enzyme histochemistry for 5-nucleotidase in presence of levamisole for lymph capills; alkaline phosphatase for blood capills. 11114A. Miura,M; Kato,S; von Ludinghausen,M (1998): Lymphatic drainage of the cerebrospinal fluid from monkey spinal meninges with special reference to the distribution of the epidural lymphatics. Arch. Histol. Cytol. 61(3, Aug), 277-286. 5'-nucleotidase staining for lymphatics; alk phosphatase for blood capillaries (Kato et al '91,'93). Also traced carbon particles from cisterna magna. Lymphatics and carbon found on surfaces of cervical & thoracic (most at brachial plexus levels) roots; not lumbosacral. Epidural lymphatics most developed on dorsal surface of lower cervical dura. I hope this helps. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Hawkins, Hal K." wrote: > > > For a model of necrotizing enterocolitis in the mouse we need to > distinguish small veins from lymphatics in the gut. Could someone > remind me of any immunostains or lectin that will allow us to tell the > two apart? > > Many thanks, > > Hal Hawkins, UTMB Galveston > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 10 Date: Thu, 06 Oct 2005 00:57:15 -0400 (EDT) From: anh2006@med.cornell.edu Subject: Re: [Histonet] lymphatics To: John Kiernan , histonet@lists.utsouthwestern.edu Message-ID: <48830.66.108.123.192.1128574635.squirrel@webmail> Content-Type: text/plain; charset=iso-8859-1 I read Dr. Kiernan's post with much interest. Has anyone on Histonet had direct experience with these protocols? I would love to get some first hand experience feedback before I try it out. Thanks, Andrea > Dear Hal Hawkins, > > The endothelium of lymphatic vessels can be stained by > enzyme activity histochemistry. The enzyme is a > 5-nucleotidase (belongs to the phosphatase family). > Ordinarily this type of method is carried out with frozen > sections of tissue that has been minimally fixed in > formaldehyde or glutaraldehyde, or with unfixed cryostat > sections that have been briefly fixed in cold acetone. The > enzyme in lymphatics may be fairly robust because its > activity can survive embedding in glycol methacrylate. > > The references below may help. If you choose one of these > methods it will be important to consult the original > publication because there's more than one sort of > 5-nucleotidase. Each ref is followed by brief notes that I > took when reading the papers. [An R in the acquisition > number means I've got a Reprint of the whole paper; an A > usually means I have a copy of the paper's Abstract. No > letter with the acquisition number usually means I've seen > the whole paper and took notes in the library.] > > 8194R. Kato,S; Yasunaga,A; Uchida,U (1991): > Enzyme-histochemical method for identification of lymphatic > capillaries. Lymphology 24, 125-129. > Glycol methacrylate-embedded sections stained for > 5'-nucleotidase & alkaline phosphatase. 5-N-ase in lymph > capills; AlkP-ase in blood capills. > > 9239R. Nishida,S; Ohkuma,M (1993): Enzyme-histochemical > staining of dermal lymphatic capillaries by guanylate > cyclase. Lymphology 26, 195-199. > Enzyme histochemical staining distinguishes lymph from > blood capillaries. Says 5-nucleotidase & adenylate cyclase > do so too. (Used unfixed cryostat sections.) > > 9665R. Okada,E (1994): An improved enzyme-histochemical > method for identification of lymphatic capillaries on > paraffin sections. Lymphology 27, Suppl, 732-735. > Staining of blood & lymph capillaries. Enzyme > histochemistry for 5-nucleotidase in presence of levamisole > for lymph capills; alkaline phosphatase for blood capills. > > 11114A. Miura,M; Kato,S; von Ludinghausen,M (1998): > Lymphatic drainage of the cerebrospinal fluid from monkey > spinal meninges with special reference to the distribution > of the epidural lymphatics. Arch. Histol. Cytol. 61(3, Aug), > 277-286. > 5'-nucleotidase staining for lymphatics; alk phosphatase > for blood capillaries (Kato et al '91,'93). Also traced > carbon particles from cisterna magna. Lymphatics and carbon > found on surfaces of cervical & thoracic (most at brachial > plexus levels) roots; not lumbosacral. Epidural lymphatics > most developed on dorsal surface of lower cervical dura. > > I hope this helps. > -- > ------------------------------- > John A. Kiernan > Department of Anatomy and Cell Biology > The University of Western Ontario > London, Canada N6A 5C1 > kiernan[AT]uwo.ca > http://publish.uwo.ca/~jkiernan/ > http://instruct.uwo.ca/anatomy/530/index.htm ------------------------------ Message: 11 Date: Thu, 6 Oct 2005 09:05:33 +0100 From: "Ahmed, T \(Tanni\)" Subject: [Histonet] Fixation using Modified Bouins and NBF To: Message-ID: <09E7875E806DFB4BBECBADBF966BDBB6F6742B@mksn79.d50.intra> Content-Type: text/plain; charset="us-ascii" Dear Histonetters, I would like to know if fixation of avian bursa tissue with Modified Bouins fixative over 10% Neutral Buffered Formalin has any advantagous effect on immuno precipitation? Thanks in advance, Tanni Tanni S Ahmed Scientific Officer - Histopathology, R&D Intervet UK Ltd. Walton Manor, Walton, Milton Keynes, Buckinghamshire, MK7 7AJ, UK. Tel. +44(0)1908 685552/685543 Fax +44(0)1908 685614 E-mail: tanni.ahmed@intervet.com -------------------------------------- This message, including attachments, is confidential and may be privileged. If you are not an intended recipient, please notify the sender then delete and destroy the original message and all copies. You should not copy, forward and/or disclose this message, in whole or in part, without permission of the sender. -------------------------------------- ------------------------------ Message: 12 Date: Thu, 6 Oct 2005 05:03:32 -0400 From: "Lee & Peggy Wenk" Subject: RE: [Histonet] 20 micron sections To: "'cynthia haynes'" , Message-ID: Content-Type: text/plain; charset="us-ascii" Several suggestions: - Cut slower (real slow; real, real slow) - Cut the blocks at room temperature (not iced/chilled). Chilled is great for getting thiner sections, warmer for thicker sections - Might have to change the knife angle. Try cutting an "empty" paraffin block (make a block with no tissue), and increase and decrease the knife angle, until you can get sections without wrinkles from the plain block of paraffin. - If all the above fail, float the section in a container of 25% alcohol, then pick the section up on a slide, drain the slide of the alcohol, and slower lower the slide almost horizontal into the warm water flotation bath. The folds need to be parallel to the surface of the water bath, not perpendicular, in order for them to be pulled out by the difference in surface tension between the water and the alcohol. If 25% alcohol is ripping the section apart when laid on the water, try a lower percent of alcohol. Let us know what worked for you, so we all can learn. Peggy A. Wenk, HTL(ASCP)SLS William Beaumont Hospital Royal Oak, MI 48073 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of cynthia haynes Sent: Wednesday, October 05, 2005 5:14 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] 20 micron sections Hello, everyone I am in need of information. I am cutting brain(human)tissue at 6 microns and 20 microns. The 6 microns sections are perfect. The 20 microns sections are an nightmare(and I mean Elm street nightmare). Can anyone suggest the best way to cut these sections without getting wrinkles and folds. I am at my wits end. Thanks in advance Cynthia Haynes H.T. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 13 Date: Thu, 6 Oct 2005 10:26:11 +0100 From: Stephen.Eyres@sanofi-aventis.com Subject: [Histonet] GFAP protocol for fluorescence To: Message-ID: Content-Type: text/plain; charset=ISO-8859-1 Hi, Does anyone have a GFAP protocol for a fluorescence method that they could share? Many thanks Steve ---------------------------------------------------------- Le pr?sent message ainsi que ses ?ventuelles pi?ces jointes est exclusivement destin? au(x) destinataire(s), personnes physiques ou morales, qu'il d?signe. Il constitue de ce fait une correspondance ? caract?re priv? et peut contenir des informations confidentielles. Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser imm?diatement l'exp?diteur par retour de courrier ?lectronique puis de le d?truire, ainsi que ses ?ventuelles pi?ces jointes, sans en conserver de copie. This message, including any attachment, is intended for the use of the individual or entity to which it is addressed. It is therefore to be considered as a private correspondence which may contain confidential information. If you are not the intended recipient, please advise the sender immediately by reply e.mail and delete this message and any attachment thereto without retaining a copy. ---------------------------------------------------------- ------------------------------ Message: 14 Date: Thu, 6 Oct 2005 04:55:39 -0500 From: "Sauer, Barb" Subject: [Histonet] staining under hoods To: Message-ID: Content-Type: text/plain; charset="iso-8859-1" Does anyone use a Labconco (or another)hood to do staining of H&E's? We recently moved into a new facility and when we moved none of our stains worked right. We have not completely ruled out the water change but we now have purchased stain(instead of making our own) and the stain is good. It would really be helpful to get feedback on who has the entire staining set up under a hood in comparison to only the xylene under the hood. We also have had the problem of a Labconco hood that we purchased that is supposed to move the height to adjust to different size people using it. It's a great concept but Labconco did not supply the ventilation tubing to make it moveable, thus we are stuck with an expensive hood that doesn't move. Any ideas???? Thanks for your help. B. Sauer Histology Department Synergy Health / St. Joesph's Hospital West Bend, WI 53095 ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you have received this email in error please notify the system manager. This footnote also confirms that this email message has been swept by MIMEsweeper for the presence of computer viruses. www.mimesweeper.com ********************************************************************** ------------------------------ Message: 15 Date: Thu, 6 Oct 2005 07:51:04 -0400 From: "Weems, Joyce" Subject: RE: [Histonet] staining under hoods To: "Sauer, Barb" , Message-ID: <83AACDB0810528418AA106F9AE9B7F7E01304DE2@sjhaexc02.sjha.org> Content-Type: text/plain; charset="iso-8859-1" Can you change the vent tubing to flexible dryer vent tubing? -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Sauer, Barb Sent: Thursday, October 06, 2005 5:56 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] staining under hoods Does anyone use a Labconco (or another)hood to do staining of H&E's? We recently moved into a new facility and when we moved none of our stains worked right. We have not completely ruled out the water change but we now have purchased stain(instead of making our own) and the stain is good. It would really be helpful to get feedback on who has the entire staining set up under a hood in comparison to only the xylene under the hood. We also have had the problem of a Labconco hood that we purchased that is supposed to move the height to adjust to different size people using it. It's a great concept but Labconco did not supply the ventilation tubing to make it moveable, thus we are stuck with an expensive hood that doesn't move. Any ideas???? Thanks for your help. B. Sauer Histology Department Synergy Health / St. Joesph's Hospital West Bend, WI 53095 ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you have received this email in error please notify the system manager. This footnote also confirms that this email message has been swept by MIMEsweeper for the presence of computer viruses. www.mimesweeper.com ********************************************************************** _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet Confidentiality Notice ** The information contained in this message may be privileged and is confidential information intended for the use of the addressee listed above. If you are neither the intended recipient nor the employee or agent responsible for delivering this message to the intended recipient, you are hereby notified that any disclosure, copying, distribution or the taking of any action in reliance on the contents of this information is strictly prohibited. If you have received this communication in error, please notify us immediately by replying to the message and deleting it from your computer. Thank you. Saint Josephs Health System, Inc. ------------------------------ Message: 16 Date: Thu, 6 Oct 2005 08:13:47 -0400 From: "Houston, Ronnie" Subject: [Histonet] amyloid in FNA smears To: histonet@lists.utsouthwestern.edu Message-ID: <530361BF03351B4CAE5270A05D3037B506B71A6E@bsrexms01.bshsir.com> Content-Type: text/plain; charset="iso-8859-1" What is the best fixation for smears from a fat-pad of a patient suspected with having amyloidosis, to get optimal preservation for demonstration of amyloid? Thanks Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com ________________________________________________________________________________________________________________________________ ________________________________________________________________________________________________________________________________ The information in this communication is intended to be confidential to the Individual(s) and/or Entity to whom it is addressed. It may contain information of a Privileged and/or Confidential nature, which is subject to Federal and/or State privacy regulations. In the event that you are not the intended recipient or the agent of the intended recipient, do not copy or use the information contained within this communication, or allow it to be read, copied or utilized in any manner, by any other person(s). Should this communication be received in error, please notify the sender immediately either by response e-mail or by phone, and permanently delete the original e-mail, attachment(s), and any copies. ------------------------------ Message: 17 Date: Thu, 06 Oct 2005 07:14:55 -0500 From: "danaspears@frontiernet.net" Subject: [Histonet] High Complexity Testing - IHC To: histonet@lists.utsouthwestern.edu Message-ID: <20051006071455.wmqs0oc8scko4gw4@webmail.frontiernet.net> Content-Type: text/plain; charset="ISO-8859-1" Hello everyone! I have been looking in the archives and am still a bit confused. I have a friend who has asked me to find out who may perform IHC in her lab... Would you say the following is correct as to who can perform IHC? BS or MT/HTL AS in science HT or MLT Prior to 24 April 1995 HS graduate AND Military training (50 wks) and classified as a Medical Laboratory Specialist OR Graduate of an accredited lab/histology program. Based on the above, I am interpreting that I currently have several techs that cannot perform IHC. Two are HT(ASCP), one HS/OJT the other GED/OJT. Neither with any college, military training, nor attended an accredited program. These two cannot perform the testing. I have two trainees, one has an associates, is an MLT, and is currently sitting for his HT. (I think he is okay, or does he have to wait to pass his HT?). The last is another trainee that has not completed her degree yet, so she has a ways to go yet. She will have her BS in May, then will sit for her HTL. Once she has her BS, she meets the requirements. Does all of this sound correct? Thanks for any and all help! ------------------------------ Message: 18 Date: Thu, 6 Oct 2005 13:53:48 +0100 From: gillian.2.brown@gsk.com Subject: [Histonet] Metachromasia of mast cells in epoxy resin sections To: histonet@lists.utsouthwestern.edu Message-ID: Content-Type: text/plain; charset=us-ascii Hi Histonetters, would any one have experience of counting granules in mouse mast cells in tissues which have been formaldehyde fixed, decalcified in a formic acid based solution, post osmicated and embedded in epoxy resin? I'm taking 1 micron sections and looking to assess the 'degranulation' status of each mast cell in terms of how many are still dark blue and how many are pink ie based on the metachromatic properties of toluidine blue. I suppose my question is will the formic acid have altered the dye binding properties so that I'll get erroneous results? I'm going to experiment with tissue that does not normally have this step but any insights now would be most welcome. Many thanks Gill Brown GlaxoSmithKline Medicines Research Centre, STEVENAGE, ------------------------------ Message: 19 Date: Thu, 6 Oct 2005 13:57:11 +0100 From: "Prior, Andrew" Subject: [Histonet] Histology Journals To: "'histonet@lists.utsouthwestern.edu'" Message-ID: Content-Type: text/plain; charset="iso-8859-1" I was wondering if anyone could recommend a good histology journal, that covers a lot of the methods and equipment. I've had a look at some journals on the web, but would appreciate some feedback from regular readers on how useful you find them. Many thanks Andrew Andrew Prior Histologist Smith & Nephew Research Centre York Science Park Heslington York, YO10 5DF UK Andrew.Prior@smith-nephew.com Confidentiality. This electronic transmission is strictly confidential to Smith & Nephew and intended solely for the addressee. It may contain information which is covered by legal, professional or other privilege. If you are not the intended addressee, or someone authorised by the intended addressee to receive transmissions on behalf of the addressee, you must not retain, disclose in any form, copy or take any action in reliance on this transmission. If you have received this transmission in error, please notify the sender as soon as possible and destroy this message. ------------------------------ Message: 20 Date: Thu, 6 Oct 2005 06:25:24 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] 20 micron sections To: cynthia haynes , Histonet@lists.utsouthwestern.edu Message-ID: <20051006132524.26581.qmail@web61220.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Try increasing the cutting angle; cutting more slow and cooling the cutting surface inmediately before the section you are going to take. cynthia haynes wrote:Hello, everyone I am in need of information. I am cutting brain(human)tissue at 6 microns and 20 microns. The 6 microns sections are perfect. The 20 microns sections are an nightmare(and I mean Elm street nightmare). Can anyone suggest the best way to cut these sections without getting wrinkles and folds. I am at my wits end. Thanks in advance Cynthia Haynes H.T. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. ------------------------------ Message: 21 Date: Thu, 6 Oct 2005 06:32:56 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] Histology Journals To: "Prior, Andrew" , "'histonet@lists.utsouthwestern.edu'" Message-ID: <20051006133256.73707.qmail@web61219.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Although it could be considered as a "biassed" opinion I would suggest for you to look at the Journal of Histotechnology from the National Society of Histotechnology. Rene J. "Prior, Andrew" wrote: I was wondering if anyone could recommend a good histology journal, that covers a lot of the methods and equipment. I've had a look at some journals on the web, but would appreciate some feedback from regular readers on how useful you find them. Many thanks Andrew Andrew Prior Histologist Smith & Nephew Research Centre York Science Park Heslington York, YO10 5DF UK Andrew.Prior@smith-nephew.com Confidentiality. This electronic transmission is strictly confidential to Smith & Nephew and intended solely for the addressee. It may contain information which is covered by legal, professional or other privilege. If you are not the intended addressee, or someone authorised by the intended addressee to receive transmissions on behalf of the addressee, you must not retain, disclose in any form, copy or take any action in reliance on this transmission. If you have received this transmission in error, please notify the sender as soon as possible and destroy this message. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. ------------------------------ Message: 22 Date: Thu, 6 Oct 2005 06:41:49 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] Nevada Society of Histotechnology To: connie grubaugh , histonet@lists.utsouthwestern.edu Message-ID: <20051006134149.25590.qmail@web61225.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Connie: Could you use this oportunity to ask the members of the Nevada Society to contribute with Tricks of the Trade to be shared by all the fellow histotechs? This could be a great help for all! Rene J. Buesa connie grubaugh wrote: The Nevada Society of HIstotechnology will be having their next seminar Oct. 28th and 29th at Sunrise Hospital Auditorium in Las Vegas Nevada. There will be a Friday evening get together with the vendors. Drinks and snacks will be provided. This will start at 5:30 pm. Saturday sign in at 7 am. Coffee and rolls will be provided by Fisher Scientific. Meeting starts at 8 am. First meeting will be Pam Marcum -Is Your Tissue Processor Fighting You? Fight Back. This will be 3 ceu"s. There will be a luncheon provided by Sakura and Cardinal Health. A business meeting will also be held at this time. Afternoon meetings. 1 pm- Sandra L. Cummings, Putting the bug in Histotechnology, A basic understanding of Microorganisms. To follow Marianne Mailhoit- Exposure control plan for Blood Borne Pathogens. There will be no charge for this meeting. Hope to see everyone there. Vegas is the best place to be on Halloween weekend. Please email me with any questions, or call 702-396-4079 or 702-938-3619. Connie Grubaugh Nevada Society of Histotechnology Striving for Excellence Despite the Odds. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! for Good Click here to donate to the Hurricane Katrina relief effort. ------------------------------ Message: 23 Date: Thu, 6 Oct 2005 09:55:39 EDT From: Debbiejsiena@aol.com Subject: [Histonet] Job Openings In Dallas, Texas To: histonet@pathology.swmed.edu Message-ID: Content-Type: text/plain; charset="US-ASCII" Good Morning All Tissue Techniques In Dallas, Texas has several HT openings along with an opening for a Lab tech. If you are the best and expect the best of Histology personnel than this is the lab for you. We have work that allows people to be creative and solve problems along with doing some rearch projects.If you would like to be part of a winning team, please contact us. Debbie Siena, cell: 520-360-3013, Fred Siena, cell: 817-228-8023, Office: 972-241-6277, Fax 972-241-4747, e-mail: tissuetech@juno.com ------------------------------ Message: 24 Date: Thu, 6 Oct 2005 08:56:14 -0500 From: "Kapoor, Sue" Subject: [Histonet] CAP question To: "'histonet@lists.utsouthwestern.edu'" Message-ID: <61E9F2400F53D5119CFC00508B44E33B02E9E1B7@khmcexch.uhsi.org> Content-Type: text/plain; charset=iso-8859-1 Hi everyone, is anyone familiar with CAP #ANP.22925 "For IHC tests that provide independent predictive/prognostic information, does the patient report include info on specimen processing, the antibody clone, and the scoring method used? (e.g., hormone receptor in breast carcinoma, HER-2/neu, EGFR). "The lab should periodically compare its patient results with published benchmarks, and also evaluate interobserver variabitlity among the pathologists in the lab". how are others handling this?? Many, many thanks in advance, Sue Kapoor, HT (ASCP) Histology Coordinator Kenosha Medical Center Kenosha, WI 262-653-5570 ------------------------------ Message: 25 Date: Thu, 06 Oct 2005 10:22:25 -0400 From: Pamela Marcum Subject: [Histonet] Region II Meeting November 4th and 5th, 2005 To: Histonet@lists.utsouthwestern.edu Message-ID: <6.1.1.1.2.20051006101724.01950860@mail.vet.upenn.edu> Content-Type: text/plain; charset="iso-8859-1"; format=flowed Region II is having a meeting!! We are including a copy of the program for your review. If anyone would like a copy of the full program with registration, cost, and directions please contact me. The meeting is at the Hilton Valley Forge Hotel in King of Prussia, PA - outside Philadelphia, PA. Best Regards, Pamela A Marcum Manager, Histology Special Procedures University of Pennsylvania School of Veterinary Medicine R.S. Reynolds Jr. CORL New Bolton Center 382 West Street Road Kennett Square, PA 19348 Phone - 610-925-6278 Fax - 610-925-8120 E-mail - pmarcum@vet.upenn.edu Friday - November 4, 2005 - Region II NSH Fall Meeting Registration 7:00AM to 8:00AM WS #1- 8:00 to 11:30AM Muscle talk! Barone WS #2- 8:00 to 11:30AM Preparing for the IHC Qualification Examination (ASCP Registry) Macrea WS #3- 8:00 to 11:30AM Unlocking The Secrets of Mohs' Grossing and Cryosectioning Praet WS #4- 8:00 to 9:30AM Chemicon's Advanced Tissue Arrayer (ATA 100), tissue Microarrays, IHC Select Reagents, and IHC Manual Select Staining for High Through-Put Screening Studies Schaub Morning Break In Exhibits - 9:30 to 10:00AM WS # 5- 10:00 to 11:30AM Confocal, Multi-Photon and Deconvolution Microscopy in Clinical Diagnosis Atkins WS #6- 10:00 to 11:30AM Microwave Tissue Fixation And Processing Frei Afternoon Workshops and Seminars WS #7- 1:00 to 2:30PM Laboratory Management: Problems and Solutions Billings WS#8- 1:00 to 2:30PM What Every Histotech Should Know About Water Macrea Afternoon Break In Exhibits - 2:30 to 3:00PM WS#9- 1:00 to 4:30PM Quality Assessment of Special Stains Micciche WS#10- 1:00 to 4:30PM From Amygdala to Arabidopsis: Why's and How's of Vibrating Blade Microscopy Freeland WS#11- 3:00 to 4:30PM Mummies: Ancient to Modern Wade WS#12- 3:00 to 4:30PM Reagent Alcohol - Can't Drink It - So What Is It? Marcum Saturday - November 5, 2005 - Region II NSH Fall Meeting Registration 7:00AM to 8:00AM WS#13- 8:00 to 9:30AM Are All Tissues Created Equal? Hughes Research vs Clinical Louro WS#14- 8:00 to 9:30AM High Profile Cases Let the Stress From igor.nasonkin <@t> jhmi.edu Fri Oct 7 12:28:45 2005 From: igor.nasonkin <@t> jhmi.edu (IGOR NASONKIN) Date: Thu Oct 13 13:19:00 2005 Subject: [Histonet] entorhinal cortex Message-ID: <6eb324e61d83.4346780d@jhmimail.jhmi.edu> wondering if somebody has a method of doing cryosections with OCT-like embedding material which does not dissolve easily in PBS-based solutions. We need to section entorhinal cortex area, and the section pieces just fall apart. Need to have something holding them together like paraffin, but prefer cryo at this time. Any ideas? Thank you! Igor From Claire.Weston <@t> umassmed.edu Wed Oct 12 18:23:42 2005 From: Claire.Weston <@t> umassmed.edu (Weston, Claire) Date: Thu Oct 13 13:19:01 2005 Subject: [Histonet] Histology position in Worcester, MA Message-ID: <254B3BAF3C954E42A7074452D80FAF0904AFB0E9@edunivmail01.ad.umassmed.edu> Research Laboratory Technician The Program in Molecular Medicine at UMass Medical School, Worcester, MA is currently recruiting a histology technician to perform medical research. Opportunity exists for a highly motivated individual to join a dynamic research laboratory that is using molecular genetic approaches to study cellular signaling mechanisms. Qualifications include a bachelor's and/or master's degree in a biological science or chemistry, working knowledge of histology techniques, strong attention to detail, and a willingness to learn. Experience with immunocytochemistry and in situ hybridization techniques is an advantage. Applicants should submit a CV to: Professor Roger Davis, Program in Molecular Medicine, UMass Medical School, 373 Plantation Street, Worcester, MA 01605. Tel. 508 856 6054 Fax. 508 856 3210 E-mail: roger.davis@umassmed.edu From mcauliff <@t> umdnj.edu Thu Oct 13 13:26:47 2005 From: mcauliff <@t> umdnj.edu (Geoff McAuliffe) Date: Thu Oct 13 13:30:11 2005 Subject: [Histonet] MOUSE BRAIN FIXATION In-Reply-To: References: Message-ID: <434EA6E7.7080101@umdnj.edu> Years ago my colleagues who worked on mouse bloood would withdraw up to 1 ml of blood from the mouse heart in a similar manner, but hitting the left ventricle does take some practice. I am a little amazed at the low volume of fixative as well, but tilting the animal so that the head is lower than the heart would help. Note that the resolution of the photos is not all that good. Geoff Favara, Cynthia (NIH/NIAID) wrote: >I have this paper and hope to try this next week. Note that the fixative is >picric acid/paraformaldehyde/gluteraldehyde. I would love to get great >fixation with 1ml and without opening the chest cavity. I will let you know! > >c > >Cynthia Favara >NIAID/NIH/RML/LPVD >903 South 4th Street >Hamilton, MT 59840 >406-363-9317 > >-----Original Message----- >From: Adam Perry [mailto:kaleid11@yahoo.com] >Sent: Thursday, October 13, 2005 8:20 AM >To: histonet@lists.utsouthwestern.edu >Subject: [Histonet] MOUSE BRAIN FIXATION > >I downloaded the technical report on this variant perfusion technique. I'm >amazed that you can get good fixation of the brain with 1 ml of fixative >pumped through the heart without clamping off descending aorta, etc. Has >anyone tried this technique (or similar variants), and what kind of results >were obtained? > >I'm planning on trying it in a week or so, because I would love to get good >brain fixation with so little time, fixative and waste production...but am >wary... > >Adam Perry > >Department of Physiology and Biophysics >University of Illinois >Chicago, IL 60612 > >There have been several inquiries regarding fixation of mouse brains >recently. > >A short technical report has just been published in BioTechniques: > >Minimally invasive method for murine brain fixation >Eichenbaum KD et al >Biotechniques 2005; 39(4): 487-500 > >After simple registration, it can be accessed at >http://www.biotechniques.com > >Ronnie Houston >Director of Anatomic Pathology >Bon Secours HealthPartners Laboratories >5801 Bremo Road >Richmond, VA 23226 >(804) 2877972 >(804) 2877906 - fax >ronnie_houston@bshsi.com > > > > >--------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > -- -- ********************************************** Geoff McAuliffe, Ph.D. Neuroscience and Cell Biology Robert Wood Johnson Medical School 675 Hoes Lane, Piscataway, NJ 08854 voice: (732)-235-4583; fax: -4029 mcauliff@umdnj.edu ********************************************** From NSEARCY <@t> swmail.sw.org Thu Oct 13 13:59:37 2005 From: NSEARCY <@t> swmail.sw.org (Nita Searcy) Date: Thu Oct 13 14:00:10 2005 Subject: [Histonet] Immunoflouresence Questions Message-ID: I have a tech that is asking these questions regarding possible "new procedures". We have a new clinician that has "new" ideas. 1. Does storing slides in refrigerator after staining aid in anyway? (I had never seen this before ; always stored in dark @ room temperature). 2. IS there a mounting media that aids in permanence? 3. To my knowledge, these slides are not permanent (no matter what you do) but will inspectors really expect to see them- even in a faded condition? (we take pictures of the kidneys) Thanks From rjbuesa <@t> yahoo.com Thu Oct 13 14:28:40 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 13 14:28:59 2005 Subject: [Histonet] Immunoflouresence Questions In-Reply-To: Message-ID: <20051013192841.98300.qmail@web61217.mail.yahoo.com> Nita: You are right, immunofluorescence procedures will fade, and fast; that is the reason behind taking photographs of the results. You may keep the slides in a refrigerator and will be able to register immunofluorescence again, but WEAKER, the next day. How long will this fluorescence will remain visible in refrigerated slides depends on the case, but cannot be predicted in general. When the pathologist that is scheduled to read our procedures is not available, we keep the slides for the next day, but that is all; no permanent solution. There is no media that will retard fading (is something inherent to the FITC conjugated antibody and has nothing to do with the medium. Probably your clinician "heard" something but evidently does not know the procedural details, or he may know something that neither you or I know. Rene J. Nita Searcy wrote: I have a tech that is asking these questions regarding possible "new procedures". We have a new clinician that has "new" ideas. 1. Does storing slides in refrigerator after staining aid in anyway? (I had never seen this before ; always stored in dark @ room temperature). 2. IS there a mounting media that aids in permanence? 3. To my knowledge, these slides are not permanent (no matter what you do) but will inspectors really expect to see them- even in a faded condition? (we take pictures of the kidneys) Thanks _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Thu Oct 13 14:36:16 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 13 14:36:30 2005 Subject: [Histonet] Bone Marrow Biopsies In-Reply-To: Message-ID: <20051013193617.80375.qmail@web61213.mail.yahoo.com> What you do at the sample site depends on what the research is all about. We used to do the following: 1- at the site we took 4 to 5 smears and fixed the core in neutral (but really neutral) buffered formalin, inside a glass container (2-3 oz.). 2- after 4-6 hours the biosy was places in EDTA and was "decalcified" for 4-6 hours and latter processed, cut, stained, etc. as usual. 3- the liquid and remanent blood contained in the vial was filtered through paper filter, placed in cassette and processed in the same way as the biopsy. 4- both decalcified biopsy and filtrate of aspirate were cut as thin as possible, except for the slides used for iron or reticulin stain, that were cut at 7 um Hope this will help! Rene J. Cheryl Crowder wrote: Hi all - I have a residen whose research will include doing bone marrow biopsies. Could someone help me with protocols for bone marrows. Vet Med here does not routinely do bone marrows and the results are spotty. What I am looking for is what you do at the point of biopsy, as make smears, fix some, etc., what fixative do you use, do you decal the BM and in what? All that "stuff". If anyone can help me, it would probably be better to e-mail me directly. Thank you, Cheryl Cheryl Crowder, BA, HTL(ASCP) Chief Technologist Anatomic Pathology Department of Pathobiological Sciences School of Veterinary Medicine Louisiana State University Skip Bertman Drive Baton Rouge, LA 70803 225-578-9734 FAX: 225-578-9720 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From tpmorken <@t> labvision.com Thu Oct 13 14:42:22 2005 From: tpmorken <@t> labvision.com (Morken, Tim - Labvision) Date: Thu Oct 13 14:42:53 2005 Subject: [Histonet] Immunoflouresence Questions Message-ID: <0556BE8AC5551E4E8AF6BB9E42509BA20328D21A@usca0082k08.labvision.apogent.com> Nita, Storage of flu0rochrome-labeled tissue slides in the fridge will retard fading for up to a couple weeks, but if you look at the slides within a day or so it is not really any better than room temperature storage in my experience. Pictures should always be taken on first viewing to get the best signal. A mounting medium containing DABCO (1,4-diazabicyclo[2.2.2]octane) will help reduce fading. You can get it from Sigma, Cat# 10981 Tim Morken Lab Vision - Neomarkers www.labvision.com Free webhosting for US State Histotechnology Societies: http://www.labvisioncorp.com/demowebsite/index.cfm -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Nita Searcy Sent: Thursday, October 13, 2005 12:00 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Immunoflouresence Questions I have a tech that is asking these questions regarding possible "new procedures". We have a new clinician that has "new" ideas. 1. Does storing slides in refrigerator after staining aid in anyway? (I had never seen this before ; always stored in dark @ room temperature). 2. IS there a mounting media that aids in permanence? 3. To my knowledge, these slides are not permanent (no matter what you do) but will inspectors really expect to see them- even in a faded condition? (we take pictures of the kidneys) Thanks _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From ana.merino-trigo <@t> wanadoo.fr Thu Oct 13 14:49:13 2005 From: ana.merino-trigo <@t> wanadoo.fr (ana.merino-trigo) Date: Thu Oct 13 14:49:35 2005 Subject: [Histonet] Immunoflouresence Questions References: <20051013192841.98300.qmail@web61217.mail.yahoo.com> Message-ID: <014201c5d02f$30723ac0$01fea8c0@BABA> Hi, I'm not really an expertise, but in my first postdoc in Australia, I was working in a Cell biology department and we were using Mowiol as mounting medium. We kept the slides in dark at 4?C, and the fluorescence was the same the day after, than the week after, even I was reviewing slides after one month on the fridge, with no a great difference. However, I was working with cell lines, using as FITC, TxRed, Alexa ... antibodies and I don't really now if tissues fades quicker. Attached the protocol used to prepare Mowiol. I do want to take the opportunity to say everyone thank you, I'm learning a lot by reading Histonet disccusions. Great work everyone. Mowiol Mounting Medium pH 8.5 Materials Hopval 5-88 4.8g glycerol 12.0g MilliQ H2O 12.0ml 0.2M Tris-HCl pH 8.5 24.0ml Tris 2.42g Tris base milliQ H2O 100ml pH to 8.5 with 6N HCl Method . Place glycerol in 50ml disposable blue cap tube . Add Hopval and stir thoroughly . Add H2O and leave for 2h at R.T. . Add tris and incubate at ~53?C until the Hopval has dissolved, stirring occassionally (takes ages) . clarify by centrifugation at 3500rpm for 30min and aliquot the supernatant into 1ml in 1.5 ml tubes . store at -20?C, stable at this temp for 12 months . once defrosted, stable at R.T. for one month Thanks Ana ----- Original Message ----- From: Rene J Buesa To: Nita Searcy ; histonet@lists.utsouthwestern.edu Sent: Thursday, October 13, 2005 9:28 PM Subject: Re: [Histonet] Immunoflouresence Questions Nita: You are right, immunofluorescence procedures will fade, and fast; that is the reason behind taking photographs of the results. You may keep the slides in a refrigerator and will be able to register immunofluorescence again, but WEAKER, the next day. How long will this fluorescence will remain visible in refrigerated slides depends on the case, but cannot be predicted in general. When the pathologist that is scheduled to read our procedures is not available, we keep the slides for the next day, but that is all; no permanent solution. There is no media that will retard fading (is something inherent to the FITC conjugated antibody and has nothing to do with the medium. Probably your clinician "heard" something but evidently does not know the procedural details, or he may know something that neither you or I know. Rene J. Nita Searcy wrote: I have a tech that is asking these questions regarding possible "new procedures". We have a new clinician that has "new" ideas. 1. Does storing slides in refrigerator after staining aid in anyway? (I had never seen this before ; always stored in dark @ room temperature). 2. IS there a mounting media that aids in permanence? 3. To my knowledge, these slides are not permanent (no matter what you do) but will inspectors really expect to see them- even in a faded condition? (we take pictures of the kidneys) Thanks _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet -- No virus found in this incoming message. Checked by AVG Anti-Virus. Version: 7.0.344 / Virus Database: 267.11.14/129 - Release Date: 11/10/2005 From scoop <@t> mail.nih.gov Thu Oct 13 15:05:17 2005 From: scoop <@t> mail.nih.gov (Sharon Cooperman) Date: Thu Oct 13 15:05:34 2005 Subject: [Histonet] spleen sectioning Message-ID: Dear Histonetites, I have been asked to prepare some FFPE spleens that are cut "along the long axis". (Apparently the standard cut for hematologists/pathologists to use?) The way a spleen looks to me, there are three possible planes to cut it in; one perpendicular to the longest dimension and two parallel to the longest dimension. Of the the two planes parallel to the longest dimension, one would be parallel to the second longest dimension and one would be perpendicular to the second longest dimension. I've been cutting my spleens parallel to the longest dimension and perpendicular to the second longest dimension, because that's where I seem to get the most information, and it also looks like what I see in my hematopathology textbook. Is that correct? I know there's probably a standard way to do it that everybody knows but I'm not a histologist, pathologist or hematologist and I'm pretty uninformed about the standard clinical ways to do spleens. I would appreciate any advice, references, etc. Thanks in advance! Sharon -- Sharon Cooperman NIH, NICHD, CBMB 301.435-8417 Building 18T, room 101 301.402-0078 fax Bethesda, MD 20892 From rjbuesa <@t> yahoo.com Thu Oct 13 15:49:54 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 13 15:50:06 2005 Subject: [Histonet] Xylene or xylene substitutes free tissue processing protocol. Message-ID: <20051013204954.69678.qmail@web61223.mail.yahoo.com> To all the Histonet subscribers: Xylene, as well as all other xylene substitutes products, are noxious substances in some way or level. Guided by the concept that it is part of the mission of any histology supervisor that of providing a safe work environment for the histotech, between 1998 and 1999 I developed a processing procedure that does not use xylene or any of its commercial substitutes. Besides being absolutely innocuous this new procedure does not require any special disposal protocol and is cheaper than the conventional and almost "universal" xylene procedure. I published this method in The Journal of Histotechnology, vol.22; no.2; pp. 143-149, 2000 and was adopted as the standard tissue procesing method of my laboratory since 26 June 1999 If you cannot find the article easily and would like to have a copy of it, you can e-mail me and I will send you an electronic version of it along with some photomicrographs. Sincerely, Rene J. Buesa --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From kcrosby <@t> cellsignal.com Fri Oct 14 06:41:15 2005 From: kcrosby <@t> cellsignal.com (Crosby, Katherine) Date: Fri Oct 14 06:41:45 2005 Subject: [Histonet] Perfusion/Dye Message-ID: Hi All, I am looking for suggestions of dyes, fluorescent or otherwise, that could be incorporated into a standard mouse perfusion procedure. Thanks in advance, Katie Katie Crosby Manager, Immunohistochemistry Cell Signaling Technology 166B Cummings Center Beverly, MA 01915 Phone: 978-867-2352 Fax: 978-867-2402 E-mail: kcrosby@cellsignal.com From b003046 <@t> nf.au.dk Fri Oct 14 06:41:34 2005 From: b003046 <@t> nf.au.dk (b003046@nf.au.dk) Date: Fri Oct 14 06:41:54 2005 Subject: [Histonet] (no subject) Message-ID: <1129290094.434f996e8ed50@webmail.nf.au.dk> Hi, im trying to get the DAKO polyclonal rabbit anti-human c-kit (CD117) to work in rats in paraffin. I have tryed using antigen retrieval in microwave and down to 1:50 dilution. Also I have tryed the C-19 c-kit from Santa Cruz and used both on paraffin and frozen tissue. It just doesnt want to work! Can anybody help? Thanks, Mette K. Hagensen Department of Cardiology B, Research Unit Aarhus University Hospital, Skejby Hospital Brendstrupgaardsvej 100 8200 Aarhus N Denmark Phone: +45 89496237 Fax: +45 89496009 Mail: b003046@nf.au.dk From mcauliff <@t> umdnj.edu Fri Oct 14 07:45:47 2005 From: mcauliff <@t> umdnj.edu (Geoff McAuliffe) Date: Fri Oct 14 07:48:54 2005 Subject: [Histonet] Perfusion/Dye In-Reply-To: References: Message-ID: <434FA87B.3000904@umdnj.edu> You can put any dye in the fixative that you want to. What are you trying to achieve? Geoff Crosby, Katherine wrote: > Hi All, > I am looking for suggestions of dyes, fluorescent or otherwise, that > could be incorporated into a standard mouse perfusion procedure. > Thanks in advance, > Katie > > > > > > Katie Crosby > Manager, Immunohistochemistry > Cell Signaling Technology > 166B Cummings Center > Beverly, MA 01915 > > Phone: 978-867-2352 > Fax: 978-867-2402 > E-mail: kcrosby@cellsignal.com > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > -- -- ********************************************** Geoff McAuliffe, Ph.D. Neuroscience and Cell Biology Robert Wood Johnson Medical School 675 Hoes Lane, Piscataway, NJ 08854 voice: (732)-235-4583; fax: -4029 mcauliff@umdnj.edu ********************************************** From scoop <@t> mail.nih.gov Fri Oct 14 08:15:02 2005 From: scoop <@t> mail.nih.gov (Sharon Cooperman) Date: Fri Oct 14 08:15:24 2005 Subject: [Histonet] spleen sectioning In-Reply-To: References: Message-ID: Sorry! Mouse spleens from transgenic mice. Thanks! Sharon >human?? > >-----Original Message----- >From: histonet-bounces@lists.utsouthwestern.edu >[mailto:histonet-bounces@lists.utsouthwestern.edu]On >Behalf Of Sharon >Cooperman >Sent: 13 October 2005 21:05 >To: histonet@lists.utsouthwestern.edu >Subject: [Histonet] spleen sectioning > > >Dear Histonetites, > >I have been asked to prepare some FFPE spleens that are cut "along >the long axis". (Apparently the standard cut for >hematologists/pathologists to use?) The way a spleen looks to me, >there are three possible planes to cut it in; one perpendicular to >the longest dimension and two parallel to the longest dimension. Of >the the two planes parallel to the longest dimension, one would be >parallel to the second longest dimension and one would be >perpendicular to the second longest dimension. I've been cutting my >spleens parallel to the longest dimension and perpendicular to the >second longest dimension, because that's where I seem to get the most >information, and it also looks like what I see in my hematopathology >textbook. Is that correct? I know there's probably a standard way >to do it that everybody knows but I'm not a histologist, pathologist >or hematologist and I'm pretty uninformed about the standard clinical >ways to do spleens. I would appreciate any advice, references, etc. >Thanks in advance! > >Sharon >-- >Sharon Cooperman >NIH, NICHD, CBMB 301.435-8417 >Building 18T, room 101 301.402-0078 fax >Bethesda, MD 20892 > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet -- Sharon Cooperman NIH, NICHD, CBMB 301.435-8417 Building 18T, room 101 301.402-0078 fax Bethesda, MD 20892 From twebster <@t> nmcinc.org Fri Oct 14 09:05:18 2005 From: twebster <@t> nmcinc.org (Tim Webster) Date: Fri Oct 14 09:05:36 2005 Subject: [Histonet] Height adjustable tables Message-ID: Hi Bonnie, We are presently ordering three adjustable work tables from a company called Bostontec. (Bob Doucette 508-574 1619) Also check out Bostontec.com for images of their installations. The tables are electrically adjustable over a wide range as we have technologists ranging from 5'2" to 6'1 in height. We looked at various vendors, but settled on Bostontec after considering pricing and design flexibility. Having said that, we haven't used the tables yet, so this is no way an endorsement blah blah blah - insert liability disclaimer here. However, they have been very easy to deal with. Have a great day and good luck. Tim Tim Webster Histology Specialist Northwestern Medical Center 133 Fairfield Street St Albans, VT 05478 (802) 524-1070 x4349 (Dept. & Voice mail) twebster@nmcinc.org Statement of Confidentiality This message and any attachments are from NMC and intended only for the addressee(s). The information contained may include privileged or other confidential information. Unauthorized review, forwarding, printing, copying or distribution is strictly prohibited. Any opinions expressed are those of the author and not those of Norhtwestern Medical Center. If you should receive this message in error, please delete it and notify the sender. Thank you. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of histonet-request@lists.utsouthwestern.edu Sent: Thursday, October 13, 2005 1:56 PM To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 23, Issue 14 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit http://lists.utsouthwestern.edu/mailman/listinfo/histonet or, via email, send a message with subject or body 'help' to histonet-request@lists.utsouthwestern.edu You can reach the person managing the list at histonet-owner@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than "Re: Contents of Histonet digest..." Today's Topics: 1. RE: use of microwaves (Pixley, Sarah (pixleysk)) 2. Re: Air Bubbles in Manual Coverslipping (Rene J Buesa) 3. RE: remove grinding debris (Rittman, Barry R) 4. slide, cassette labelers (Osborn, Sharon) 5. RE: slide, cassette labelers (Schumacher, Jennifer J) 6. Re: An old tissue processor (rchiovetti@aol.com) 7. Coverslipping technics (Osborn, Sharon) 8. Re: RE: use of microwaves (Rene J Buesa) 9. Re: Cytologies crossing state lines (Rene J Buesa) 10. mounting media for EGFP in liver (Caroline Bass) 11. RE: Cytologies crossing state lines (Ford Royer) 12. decalcifying solutions for skeletochronology (Malcolm McCallum) 13. Re: decalcifying solutions for skeletochronology (John Kiernan) 14. Histotech Opportunites in your Area (Eric Dye (ext 223)) 15. Height Adjustable tables (Bonnie J. McMahill) 16. SGLT1 antibody (Jonathan Wilson) 17. RE: beta-2- microglobulin (Edwards, R.E.) 18. Re: decalcifying solutions for skeletochronology (Rene J Buesa) 19. MOUSE BRAIN FIXATION (Adam Perry) 20. RE: MOUSE BRAIN FIXATION (Favara, Cynthia (NIH/NIAID)) 21. Re: Tissue with Fat (Arshia Mian) ---------------------------------------------------------------------- Message: 1 Date: Wed, 12 Oct 2005 13:20:52 -0400 From: "Pixley, Sarah \(pixleysk\)" Subject: [Histonet] RE: use of microwaves To: Message-ID: Content-Type: text/plain; charset="us-ascii" Dear All: I wondered if anyone that is currently using microwaves to heat samples has considered using a sand dry-bath for heating? The molecular biologists have all switched to those and they are really great. They are just heaters with a well that you put a good quality sand in. They can heat the sand up to 130 degrees centigrade. Then you stick your sample in the sand. The heat is very consistent. You can have something go to close to boiling almost instantly, without the muss and fuss of a hot water bath (or a microwave). Aside from a few bits of sand getting out, they are very civilized and clean. And they take up FAR less counter space than a microwave. Just a thought, but it is from someone who is not doing HIER. Sarah ------------------------------ Message: 2 Date: Wed, 12 Oct 2005 10:22:58 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] Air Bubbles in Manual Coverslipping To: Travis Troyer , histonet@lists.utsouthwestern.edu Message-ID: <20051012172258.72021.qmail@web61225.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 You should start with the fluidity of the "glue" (I always prefer to call it "mounting medium"). It should be fluid enough to spread over the section. You should apply a very small drop over the section with either a plastic dropper or a wood applicator. The coverslip should be over a paper towel and you should get the slide/section with the mounting medium drop on it in contact with the coverslip and press it gently. You should wrap your finger with a cotton fiber disposable gauze to immediately clean the excess mounting medium. If the mounting medium is liquid enough you will prevent the bubbles, and even if the amount is in excess you can clean it around the coverslip eliminating the "overflow". Rest the coverslipped slide flat to dry out. Dammar resin or any other commercial mounting medium will do. Regardless if the diluent is toluene or xylene, you can always adjust the fluidity with xylene (which is totally miscible with toluene). Practice is essential ("like getting to Carnegie Hall!"). Hope this will help. Rene J. Travis Troyer wrote: We have recently come under the "gun" for excess bubbles and glue on the slides after coverslipping. We currently do all of coverslipping manually and was wondering if anyone had any helpful hints to keep the bubbles out and and excess glue off the slide. Thanks for the assistance, Travis Troyer Peterson Laboratory Services _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. ------------------------------ Message: 3 Date: Wed, 12 Oct 2005 13:18:31 -0500 From: "Rittman, Barry R" Subject: RE: [Histonet] remove grinding debris To: "histonet" Message-ID: Content-Type: text/plain; charset="us-ascii" Myriam. Are you using water as a lubricant on the paper? If so remember that Epon is somewhat hygroscopic. Can grind using a neutral oil. If grinding dry then repeat the grinding on a frosted glass plate. This often will remove a lot of the debris. Can first try a commercial frosted slide or prepare your own using alumina. Are you sure that the debris is not being introduced during staining? Hydroxyapatite and other minerals can cause precipitates and also may bind to some dyes. Barry -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Myri37@aol.com Sent: Wednesday, October 12, 2005 10:56 AM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] remove grinding debris Hello everyone, I grind on abrasive sandpaper, sections of titanium implants embedded in epon. Titanium is coated with a hydroxyapatite layer and soft tissue. After staining with RBS (rapid bone stain), i see on slides a lot of debris,i think they are grinding debris and titanium or hydroxyapatite dust... Does anyone have a process to remove these debris ? Do you recommend a wash with a specific detergent solution after grinding, does 0.1 % Zephiran chloride solution efficient ? Thank you very much for any advice. Myriam NI France _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 4 Date: Wed, 12 Oct 2005 14:49:25 -0400 From: "Osborn, Sharon" Subject: [Histonet] slide, cassette labelers To: "'histonet@lists.utsouthwestern.edu'" Message-ID: <29B25753F6B1D51196110002A589D44402398230@PALMSG30.us.schp.com> Content-Type: text/plain Jill, Please consider the Leica cassette and slide printers (not labelers). We have had ours almost 1 year with multiple users. They are very good, consistent and the service has been great. They are just about ready for a major upgrade to solve some problems that developed this past year with units out in the field. Still with those problems, these are better than the previous ones. As you, I have used the ShurMark for years...it was great for its time and definitely filled a void. However, the Leica is the new generation of labelers and well worth your having it demonstrated in your facility. Sharon Osborn DNAX, Schering Plough BioPharma Palo Alto, CA 650.496.6539 Date: Tue, 11 Oct 2005 12:23:42 -0700 (PDT) From: Jill Cox Subject: [Histonet] Cassette Labeler To: histonet@lists.utsouthwestern.edu Message-ID: <20051011192343.43419.qmail@web52111.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Hello Histonetters, We will be purchasing a cassette labeler soon and hopefully slide labeler down the road. I would love your feedback on which one's are more reliable. I have used a Shurmark in the past and it seemed to work fine. If there is something else out there you are using I would like to look into that as well. Thank you in advance, Jill Jill Cox HT (ASCP) ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. ------------------------------ Message: 5 Date: Wed, 12 Oct 2005 14:02:22 -0500 From: "Schumacher, Jennifer J" Subject: RE: [Histonet] slide, cassette labelers To: "Osborn, Sharon" , Message-ID: Content-Type: text/plain; charset="us-ascii" I agree completely. Our slide labeler was down one day, and we tried to use our old ShurMark as a backup. Only then did we remember how lucky we were to have the new units. They are cleaner, easier to use, and oh so FAST!!! Plus, we are saving a fortune by not buying the "colormark" slides any longer. I would highly recommend them. Jennifer Schumacher, University of Minnesota Medical Center, Fairview -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Osborn, Sharon Sent: Wednesday, October 12, 2005 1:49 PM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] slide, cassette labelers Jill, Please consider the Leica cassette and slide printers (not labelers). We have had ours almost 1 year with multiple users. They are very good, consistent and the service has been great. They are just about ready for a major upgrade to solve some problems that developed this past year with units out in the field. Still with those problems, these are better than the previous ones. As you, I have used the ShurMark for years...it was great for its time and definitely filled a void. However, the Leica is the new generation of labelers and well worth your having it demonstrated in your facility. Sharon Osborn DNAX, Schering Plough BioPharma Palo Alto, CA 650.496.6539 Date: Tue, 11 Oct 2005 12:23:42 -0700 (PDT) From: Jill Cox Subject: [Histonet] Cassette Labeler To: histonet@lists.utsouthwestern.edu Message-ID: <20051011192343.43419.qmail@web52111.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Hello Histonetters, We will be purchasing a cassette labeler soon and hopefully slide labeler down the road. I would love your feedback on which one's are more reliable. I have used a Shurmark in the past and it seemed to work fine. If there is something else out there you are using I would like to look into that as well. Thank you in advance, Jill Jill Cox HT (ASCP) ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 6 Date: Wed, 12 Oct 2005 15:03:44 -0400 From: rchiovetti@aol.com Subject: Re: [Histonet] An old tissue processor To: pruegg@ihctech.net, histonet@pathology.swmed.edu Message-ID: <8C79D76CD48E79F-17C4-3A40@MBLK-M07.sysops.aol.com> Content-Type: text/plain; charset="us-ascii" Patsy, Boy that *is* an oldie! The Fisher products were taken over by RMC, Inc. several years ago. Then RMC sold the product line to Ventana Medical Systems here in Tucson. It's a long shot, but you might try calling Ventana at (800) 227-2155. Ask for Tom Kennedy. Tom's was with the product line from the original Fisher days, then RMC, then Ventana. He would know if anyone would about parts for the 166. Good Luck! Cheers, Bob Chiovetti The Microscope works Arizona's Microscopy Resource Tucson, Arizona USA Tel./Fax 520-546-4986 Member, Arizona Small Business Association (www.asba.com) -----Original Message----- From: Patsy Ruegg To: histonet@pathology.swmed.edu Sent: Wed, 12 Oct 2005 10:03:36 -0600 Subject: [Histonet] An old tissue processor We have an old Fisher Histomatic Model 166A tissue processor that is on the fritz. Does anyone know where we might be able to find parts and service for such a critter???? Best regards, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 7 Date: Wed, 12 Oct 2005 15:03:52 -0400 From: "Osborn, Sharon" Subject: [Histonet] Coverslipping technics To: "'histonet@lists.utsouthwestern.edu'" Message-ID: <29B25753F6B1D51196110002A589D44402398231@PALMSG30.us.schp.com> Content-Type: text/plain Travis, One of my favorite tasks is coverslipping. However, due to pressing duties and volume, the automated coverslipper does the work. To get a good coverslip technic is essential when doing so manually. This is the one I use: 1. The slide is moist from the xylene. I use a dropper (I do have one of the old type bottles with the little glass dropper with a round glass bulb on its end)or a wooden applicator stick. Dip the stick into xylene to moisten it then into the mounting medium. The mounting medium will form a ball drop on the applicator. 2. Place the ball drop of mounting medium directly on the lower edge of the slide. I generally make a little line of it along the edge. 3. Pick up the coverslip with two fingers(I use thumb and index) along the coverslip edges. Be certain there are no sticky coverslips stuck together. Place the long edge of the coverslip along the lower edge of the slide on the mounting medium. The slide will gently lay down on the mounting medium and move it over the slide as it makes contact. There are rarely any bubbles. 4. If there are bubbles, gentle pressure with forceps or the applicator end (not one with mounting medium) will move those out. Also, there is rarely any excell medium around the edges. I 5. If there is excess mountimg medium around the edges, wipe it gently with a Kimwipe moistened with xylene or use a #3 artist brush (camel hair) dipped in xylene to clean the edges of the slide. Lay in mats to dry. Practise will develop this technic such that you will be faster than the automated coverslipper--the glass ones--which are all I recommend!..:-) Sharon Osborn DNAX, Schering Plough BioPharma Palo Alto, CA 650.496.6539 Date: Wed, 12 Oct 2005 11:58:11 -0500 From: "Travis Troyer" Subject: [Histonet] Air Bubbles in Manual Coverslipping To: Message-ID: <003501c5cf4e$217bb000$6601010a@Peterson.local> Content-Type: text/plain; charset="iso-8859-1" We have recently come under the "gun" for excess bubbles and glue on the slides after coverslipping. We currently do all of coverslipping manually and was wondering if anyone had any helpful hints to keep the bubbles out and and excess glue off the slide. Thanks for the assistance, Travis Troyer Peterson Laboratory Services ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. ------------------------------ Message: 8 Date: Wed, 12 Oct 2005 13:45:52 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] RE: use of microwaves To: "Pixley, Sarah \(pixleysk\)" , histonet@lists.utsouthwestern.edu Message-ID: <20051012204552.15123.qmail@web61225.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 I have used sand bath during some biochemistry procedures and I agree with you that they are great BUT (there is always a BUT) temperature regulation is somewhat tricky and will depend on the amount of sand and the condition of the heating elements. For HIER you will have to consider the following aspects: 1- it requires almost one half an hour to complete. 2- How much HIER solution are you going to use? If not enough it will probably evaporate while in the sand bath after just a few minutes. 3- evaporation is prevented in the steamer because the environment is water vapor saturated and the vapor phase is, lets say "compensated". 4- in the microwave oven you will limit evaporation because HIER is quicker (due to the effect of the microwaves on the bipolar molecules within the tissue, and this, along with heat, is the fundament for the microwave oven). You will have to do many tests to find out an ideal ratio volume/temperatureXtime for a HIER done in a sand bath. The steamer is so simple to operate, and so consistent, that I personally would not try the sand bath to do HIER. This is just my opinion. Hope this will help! Rene J. "Pixley, Sarah (pixleysk)" wrote: Dear All: I wondered if anyone that is currently using microwaves to heat samples has considered using a sand dry-bath for heating? The molecular biologists have all switched to those and they are really great. They are just heaters with a well that you put a good quality sand in. They can heat the sand up to 130 degrees centigrade. Then you stick your sample in the sand. The heat is very consistent. You can have something go to close to boiling almost instantly, without the muss and fuss of a hot water bath (or a microwave). Aside from a few bits of sand getting out, they are very civilized and clean. And they take up FAR less counter space than a microwave. Just a thought, but it is from someone who is not doing HIER. Sarah _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. ------------------------------ Message: 9 Date: Wed, 12 Oct 2005 13:49:14 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] Cytologies crossing state lines To: "Michelle D. Moore" , Histonet Message-ID: <20051012204914.21514.qmail@web61212.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 I would contact UPS or FederalExpress. Some time ago we had to send some samples and they knew all the regulations/limitations (since they are the carriers). Rene J. "Michelle D. Moore" wrote: Hello in histoland. I am looking for information on whether or not you can ship freshly collected pap smears across state lines?! I have been told you can by one place and then told that you cannot at all. I know that you cannot send them across California's state line but what about the rest of the nation? I appreciate any help I can get. Thank you for your time and help. Michelle D. Moore The Memorial Hospital Craig, CO tmhpath@amigo.net _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. ------------------------------ Message: 10 Date: Wed, 12 Oct 2005 17:19:07 -0400 From: Caroline Bass Subject: [Histonet] mounting media for EGFP in liver To: Histonet (E-mail) Message-ID: <41489ADD-7323-463D-88F8-42BBAC1B341A@bidmc.harvard.edu> Content-Type: text/plain; charset=US-ASCII; delsp=yes; format=flowed Hello, I have used a viral vector to transduce liver and pancreatic tissues in the mouse with EGFP and RFP. I have cut portions of the liver and pancreas into small blocks and fixed overnight with NBF. What is the best way to visualize the signal? I want to cut 30 micron sections, as I only have access to a sliding microtome. Could someone recommend a good, and hopefully inexpensive, mounting media for fluorescence? I have a fairly strong signal but I am worried about losing it if I use the wrong product. I am not very experienced with fluorescence so I really don't know what to use. Someone recommended Aqua PolyMount from polysciences. Any suggestions would be appreciated. Caroline ------------------------------ Message: 11 Date: Wed, 12 Oct 2005 16:19:09 -0500 From: "Ford Royer" Subject: RE: [Histonet] Cytologies crossing state lines To: "Michelle D. Moore" , "Histonet" Message-ID: Content-Type: text/plain; charset="iso-8859-1" I believe that it is okay, as long as it is not for an immoral purpose... ...sorry, couldn't resist. ;-) (For you youngsters out there... see "The Mann Act of 1910") ~ Ford Ford Royer, MT(ASCP) Minnesota Medical Specialists 7177 Madison Ave. W. Golden Valley, MN 55427-3601 888-790-9686 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Michelle D. Moore Sent: Wednesday, October 12, 2005 11:50 AM To: Histonet Subject: [Histonet] Cytologies crossing state lines Hello in histoland. I am looking for information on whether or not you can ship freshly collected pap smears across state lines?! I have been told you can by one place and then told that you cannot at all. I know that you cannot send them across California's state line but what about the rest of the nation? I appreciate any help I can get. Thank you for your time and help. Michelle D. Moore The Memorial Hospital Craig, CO tmhpath@amigo.net _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 12 Date: Wed, 12 Oct 2005 23:01:16 -0500 From: "Malcolm McCallum" Subject: [Histonet] decalcifying solutions for skeletochronology To: Message-ID: Content-Type: text/plain; charset="iso-8859-1" Hi, I am having some students do various skeletochronology projects with amphibians and reptiles. I could just buy some decalcifying solution, but we have so many acids, I figured I would make some. Any good recipes out there for decalcifying bone w/o heating? Malcolm L. McCallum Assistant Professor Department of Biological Sciences Texas A&M University Texarkana 2600 Robison Rd. Texarkana, TX 75501 O: 1-903-233-3134 H: 1-903-791-3843 Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html ------------------------------ Message: 13 Date: Thu, 13 Oct 2005 00:57:47 -0400 From: John Kiernan Subject: Re: [Histonet] decalcifying solutions for skeletochronology To: Malcolm McCallum Cc: histonet@lists.utsouthwestern.edu Message-ID: <434DE94B.61DC3E1C@uwo.ca> Content-Type: text/plain; charset=us-ascii Dear Dr McCallum, Every book in the field of histotechnology contains several recipes for decalcifying. The calcified material can be removed by dissolving in a suitable acid (usually hydrochloric or formic) or by chelation with a suitable anion (usually citrate or EDTA). The choice of method is based on the requirements of the investigation. If you are in doubt about choosing a method, ask the internet for references rather than recipes. There are scores of books and hundreds of papers! A solid place to start searxhing the literature is Pearse's Histochemistry. John A. Kiernan MB, ChB, PhD, DSc Professor, Dept of Anatomy & Cell Biology The University of Western Ontario London, Canada. ---------------------------- Malcolm McCallum wrote: > > Hi, > I am having some students do various skeletochronology projects with amphibians and reptiles. I could just buy some decalcifying solution, but we have so many acids, I figured I would make some. Any good recipes out there for decalcifying bone w/o heating? > > Malcolm L. McCallum > Assistant Professor > Department of Biological Sciences > Texas A&M University Texarkana > 2600 Robison Rd. > Texarkana, TX 75501 > O: 1-903-233-3134 > H: 1-903-791-3843 > Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html > ------------------------------ Message: 14 Date: Wed, 12 Oct 2005 20:58:54 -0400 From: Eric Dye (ext 223) Subject: [Histonet] Histotech Opportunites in your Area To: Histonetters Message-ID: Content-Type: text/plain Histonetters I'm presently on a search for my best client companies seeking HistoTech Supervisors and Histo Bench Techs. These are mostly fulltime permanent positions. (I have a few travel/temp HistoTech positions as well - see below). Most of my Histo Techs positions are clinical, although I do have a few Research Histo Tech positions as well. Here are some of my Hottest jobs: 1. Louisiana( North Western) Openings for a HistoTech(Bench) No weekends, No call, and Top Dollar. 2. Ohio(Dayton Area) HistoTech Manager No Weekends, No call, Top Dollar if you are interested, please call me today at 1-800-466-9919 ext 223 3. Colorado (Greater Denver area) Opening for Several Bench Positions(Night Shift) No weekends, No call, Top Dollar 4. New Hampshire Openings for several Histotechs(Bench) No Weekends, No Call, Top Dollar, Excellent Benefits if you are interested, please call me today at 1-800-466-9919 ext 223 5. Minnesota Opening for HistoTech(Bench) No weekends, No Call, Top Dollar 6. New Jersey(Southern) Openings for HistoTech(Bench) No Weekends, No Call, Top Dollar if you are interested, please call me today at 1-800-466-9919 ext 223 7.Rhode Island Openings for Histotech(Bench) No weekends, No call, Top Dollar 8. California( Southern) HistoTech Supervisor(2nd shift) Openings for Histotechs(Bench,2nd shift) Great Location, No weekends, No call, Top Dollar 9. California(Southern) Openings for HistoTechs(Bench) Great Location, No weekends, No call, Top Dollar 10. Pennsylvania(Multiple jobs in greater Pittsburgh area) HistoTech opening(Bench) if you are interested, please call me today at 1-800-466-9919 ext 223 11. Illinois(Multiple jobs in Greater Illinois area) Opening for Histotech(Bench) 12. Michigan(Multiple jobs in Greater Michigan area) Openings for Histotechs(Bench) 13. Indiana Opening for Histotech(Bench) 14. Washington State(Eastern) Opening for Lead HistoTech(Bench) No weekends, No call if you are interested, please call me today at 1-800-466-9919 ext 223 15. Oklahoma Opening for Histotech(Bench) No weekends, No call. 16. Massachusetts (Boston Area) Part time Histo Tech(Permanent) No weekends, No call 17. Tennesee(Memphis area) Openings for Supervisor and Bench Techs 18. New Mexico openings for Supervisor and Bench Techs No Weekends, No Call, Excellent Benefits.. 19. Nebraska Openings for Histo Techs(Bench) No Weekends, No Call, Top Dollar 20. Virginia(Western Virginia) Opening for Bench Histotech No weekends, No call if you are interested, please call me today at 1-800-466-9919 ext 223 21. Ohio( Southern) Opening for Bench HistoTech No weekends, No call 22. Maryland( Baltimore area) Opening for Bench Histotech 23. Missouri(Greater Missouri area) Opening for Bench Histotech No weekends, No call 24. Wisconsin(Eastern area) Opening for a Bench Histotech No weekends, No call 25. Florida(Greater Florida area) Openings for Bench Histotech No weekends, No call The clients are currently interviewing candidates - so if you are interested, please call me today at 1-800-466-9919 ext 223 Thank you, Eric Dye-Sr Allied Healthcare Recruiter 1-800-466-9919 ext 223 ------------------------------ Message: 15 Date: Wed, 12 Oct 2005 23:06:56 -0700 From: "Bonnie J. McMahill" Subject: [Histonet] Height Adjustable tables To: Message-ID: Content-Type: text/plain; charset="utf-8" Hi all, I was wondering if anyone has height adjustable (electronic or crank) tables for cutting? We are putting some in our lab, but are interested in purchasing some from a company that has a known track record. Thanks! Bonnie McMahill InCyte Pathology Spokane, WA ------------------------------ Message: 16 Date: Thu, 13 Oct 2005 11:25:37 +0100 From: "Jonathan Wilson" Subject: [Histonet] SGLT1 antibody To: Message-ID: <003101c5cfe0$751b8950$17dea8c0@ciimar.up.pt> Content-Type: text/plain; format=flowed; charset="iso-8859-1"; reply-type=original Hello, Could someone please recommend an antibody against SGLT1 (sodium glucose transporter) that works in rat or mouse FFPE tissue. I have tried a chemicon antibody but without any luck. Thanks for any advice. Sincerely, Jonathan Wilson CIIMAR-Ecofisiologia Rua dos Bragas 289 4050-123 Porto Portugal tel 351 22 340 1809 fax 351 22 339 0608 alt. e-mail: mop01258@mail.telepac.pt ------------------------------ Message: 17 Date: Thu, 13 Oct 2005 12:27:10 +0100 From: "Edwards, R.E." Subject: [Histonet] RE: beta-2- microglobulin To: Message-ID: Content-Type: text/plain; charset="iso-8859-1" Looking for an antibody to the above that works, as ever, on paraffin processed mouse tissues... Many thanks Richard Edwards MRC TOX UNIT LEICESTER...U.K.... ------------------------------ Message: 18 Date: Thu, 13 Oct 2005 07:48:24 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] decalcifying solutions for skeletochronology To: Malcolm McCallum , histonet@lists.utsouthwestern.edu Message-ID: <20051013144824.30771.qmail@web61214.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 The decalcifying solution that you may end using (after any standard recipe you can find in any good histology manual) will depend on the type of bone you want to decalcify and what details you want to preserve. EDTA will give you the most details, but it will take longer; sulfuric acid will decalcify faster but will destroy almost any detail. The weaker the acid (lique citric) the slower the process but the more detail you will preserve; and an inverse correlation you will find with the stronger the acid. I hope this will help you to sort out your options. Rene J. Malcolm McCallum wrote: Hi, I am having some students do various skeletochronology projects with amphibians and reptiles. I could just buy some decalcifying solution, but we have so many acids, I figured I would make some. Any good recipes out there for decalcifying bone w/o heating? Malcolm L. McCallum Assistant Professor Department of Biological Sciences Texas A&M University Texarkana 2600 Robison Rd. Texarkana, TX 75501 O: 1-903-233-3134 H: 1-903-791-3843 Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. ------------------------------ Message: 19 Date: Thu, 13 Oct 2005 08:19:46 -0700 (PDT) From: Adam Perry Subject: [Histonet] MOUSE BRAIN FIXATION To: histonet@lists.utsouthwestern.edu Message-ID: <20051013151946.35658.qmail@web30411.mail.mud.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 I downloaded the technical report on this variant perfusion technique. I'm amazed that you can get good fixation of the brain with 1 ml of fixative pumped through the heart without clamping off descending aorta, etc. Has anyone tried this technique (or similar variants), and what kind of results were obtained? I'm planning on trying it in a week or so, because I would love to get good brain fixation with so little time, fixative and waste production...but am wary... Adam Perry Department of Physiology and Biophysics University of Illinois Chicago, IL 60612 There have been several inquiries regarding fixation of mouse brains recently. A short technical report has just been published in BioTechniques: Minimally invasive method for murine brain fixation Eichenbaum KD et al Biotechniques 2005; 39(4): 487-500 After simple registration, it can be accessed at http://www.biotechniques.com Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. ------------------------------ Message: 20 Date: Thu, 13 Oct 2005 11:35:32 -0400 From: "Favara, Cynthia (NIH/NIAID)" Subject: RE: [Histonet] MOUSE BRAIN FIXATION To: 'Adam Perry' , histonet@lists.utsouthwestern.edu Message-ID: Content-Type: text/plain I have this paper and hope to try this next week. Note that the fixative is picric acid/paraformaldehyde/gluteraldehyde. I would love to get great fixation with 1ml and without opening the chest cavity. I will let you know! c Cynthia Favara NIAID/NIH/RML/LPVD 903 South 4th Street Hamilton, MT 59840 406-363-9317 Disclaimer: The information in this e-mail and any of its attachments is confidential and may contain sensitive information. It should not be used by anyone who is not the original intended recipient. If you have received this e-mail in error please inform the sender and delete it from your mailbox or any other storage devices. National Institute of Allergy and Infectious Diseases shall not accept liability for any statements made that are sender's own and not expressly made on behalf of the NIAID by one of its representatives -----Original Message----- From: Adam Perry [mailto:kaleid11@yahoo.com] Sent: Thursday, October 13, 2005 8:20 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] MOUSE BRAIN FIXATION I downloaded the technical report on this variant perfusion technique. I'm amazed that you can get good fixation of the brain with 1 ml of fixative pumped through the heart without clamping off descending aorta, etc. Has anyone tried this technique (or similar variants), and what kind of results were obtained? I'm planning on trying it in a week or so, because I would love to get good brain fixation with so little time, fixative and waste production...but am wary... Adam Perry Department of Physiology and Biophysics University of Illinois Chicago, IL 60612 There have been several inquiries regarding fixation of mouse brains recently. A short technical report has just been published in BioTechniques: Minimally invasive method for murine brain fixation Eichenbaum KD et al Biotechniques 2005; 39(4): 487-500 After simple registration, it can be accessed at http://www.biotechniques.com Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ Message: 21 Date: Thu, 13 Oct 2005 09:32:24 -0700 From: "Arshia Mian" Subject: [Histonet] Re: Tissue with Fat To: Message-ID: Content-Type: text/plain; charset="us-ascii" I have been having some problems with cutting Tissue with Fat on a cryostat (box temp and object temp are at -20C). The slices I cut tend to curl in the middle of the block close to the tissue. I have changed the blade several times, but it still continues to happen even with a new blade. Is there any correlation between the box temp and the object temp that would cause this to happen? Any suggestions? I have also noticed that at times, even though my setting is set to 20 microns, that I get thick and thin cuts. Any suggestions on this as well? I would greatly appreciate it. Thanks. ------------------------------ _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet End of Histonet Digest, Vol 23, Issue 14 **************************************** From fladanielski <@t> yahoo.com.br Fri Oct 14 09:12:53 2005 From: fladanielski <@t> yahoo.com.br (Flavia Danielski) Date: Fri Oct 14 09:13:23 2005 Subject: [Histonet] 6um pancreas slice Message-ID: <20051014141253.34096.qmail@web30607.mail.mud.yahoo.com> histo-people: I would be really glad if someone could help me with a basic protocol for cryosections of pancreas... my goal is to reach 6um sections... does OCT work in this case? Tks Flavia Fl?via Helena da Silva Lab. de Imunogen?tica, Depto. de Gen?tica, UFRGS Av. Bento Gon?alves, 9500 pr?dio 43323/lab 212C Bairro Agronomia. POA-RS-BRAZIL phone: 0055-(51)33166737 _______________________________________________________ Promo??o Yahoo! Acesso Gr?tis: a cada hora navegada voc? acumula cupons e concorre a mais de 500 pr?mios! Participe! http://yahoo.fbiz.com.br/ From Dorothy.L.Webb <@t> HealthPartners.Com Fri Oct 14 09:20:19 2005 From: Dorothy.L.Webb <@t> HealthPartners.Com (Webb, Dorothy L) Date: Fri Oct 14 09:23:08 2005 Subject: [Histonet] Dry tissue Message-ID: <0E394B648E5284478A6CCB78E5AFDA2718C46B@hpes1.HealthPartners.int> I thought I knew most of the processing answers when hit with problematic tissues to cut, but, lately, I am stymied!! Lately, our smaller biopsies have been very dry and nothing has changed in our processing routine or reagents. I did lessen the times in absolute a little and also let the time in xylene be lessened. I am not seeing the change I thought I would. Can anyone give me any suggestions as to how to process small biopsies in the same processor with large specimens without drying out the small GI biopsies and still getting the proper processing on large speicmens. Thanks ahead of time for responding to this!! ________________________________________ This e-mail and any files transmitted with it are confidential and are intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient or the individual responsible for delivering the e-mail to the intended recipient, please be advised that you have received this e-mail in error and that any use, dissemination, forwarding, printing, or copying of this e-mail is strictly prohibited. If you have received this e-mail in error, please immediately notify the HealthPartners Support Center by telephone at (952) 967-6600. You will be reimbursed for reasonable costs incurred in notifying us. From Wanda.Smith <@t> HCAhealthcare.com Fri Oct 14 09:31:27 2005 From: Wanda.Smith <@t> HCAhealthcare.com (Smith Wanda) Date: Fri Oct 14 09:31:41 2005 Subject: [Histonet] Looking for PRN Histotech Message-ID: Good Morning and Happy Friday, I am trying to fill a PRN/fill-in Histotech position in the Charleston, South Carolina area in a fun lab to work in. Hours will vary, however it may be one 8 hour day per week and fill-in/on call when a tech calls in sick. Please contact me by email, phone or fax with questions or consideration. Thanks, Wanda Wanda G. Smith, HTL/HT(ASCP) Pathology Supervisor Trident Medical Center 9330 Medical Plaza Drive North Charleston, SC 29406 843-797-4586 fax 843-797-4296 wanda.smith@hcahealthcare.com From abright <@t> brightinstruments.com Fri Oct 14 09:56:46 2005 From: abright <@t> brightinstruments.com (Alan Bright) Date: Fri Oct 14 09:50:47 2005 Subject: [Histonet] 6um pancreas slice Message-ID: Dear Flavia, I would ideally use an embedding medium, and section with the specimen temperature at -10? to -12?C and the knife temperature at -25?C or below. Best Regards Alan Bright Bright Instrument Co.Ltd. St Margaret's Way Huntingdon Cambridgeshire PE29 6EU England Tel No:+44 (0)1480 454528 Fax No:+44 (0)1480 456031 Email: abright@brightinstruments.com Web Site: www.brightinstruments.com -----Original Message----- From: Flavia Danielski [mailto:fladanielski@yahoo.com.br] Sent: 14 October 2005 15:13 To: lista discussao Subject: [Histonet] 6um pancreas slice histo-people: I would be really glad if someone could help me with a basic protocol for cryosections of pancreas... my goal is to reach 6um sections... does OCT work in this case? Tks Flavia Fl?via Helena da Silva Lab. de Imunogen?tica, Depto. de Gen?tica, UFRGS Av. Bento Gon?alves, 9500 pr?dio 43323/lab 212C Bairro Agronomia. POA-RS-BRAZIL phone: 0055-(51)33166737 _______________________________________________________ Promo??o Yahoo! Acesso Gr?tis: a cada hora navegada voc? acumula cupons e concorre a mais de 500 pr?mios! Participe! http://yahoo.fbiz.com.br/ _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From PMonfils <@t> Lifespan.org Fri Oct 14 09:51:39 2005 From: PMonfils <@t> Lifespan.org (Monfils, Paul) Date: Fri Oct 14 09:51:54 2005 Subject: [Histonet] spleen sectioning Message-ID: <09C945920A6B654199F7A58A1D7D1FDE017175F3@lsexch.lsmaster.lifespan.org> Sharon, The way you are sectioning them is the standard way. Because of the natural curvature of a mouse spleen, sectioning them parallel to the long axis and parallel to the second longest dimension often makes it impossible to get the full length of the spleen in one section, and it also creates low angle tangential sections of the capsule at one or both ends. Sections perpendicular to the long axis are distortion free, but of course represent only one point along the length of the spleen. The way you are cutting them, parellel to the long axis and perpendicular to the second longest dimension, gives a full representation of the entire length of the spleen, with perpendicular sections of the capsule all the way around. It's definitely the preferred way. > ---------- > From: histonet-bounces@lists.utsouthwestern.edu on behalf of > Sharon Cooperman > Sent: Thursday, October 13, 2005 1:05 PM > To: histonet@lists.utsouthwestern.edu > Subject: [Histonet] spleen sectioning > > Dear Histonetites, > > I have been asked to prepare some FFPE spleens that are cut "along > the long axis". (Apparently the standard cut for > hematologists/pathologists to use?) The way a spleen looks to me, > there are three possible planes to cut it in; one perpendicular to > the longest dimension and two parallel to the longest dimension. Of > the the two planes parallel to the longest dimension, one would be > parallel to the second longest dimension and one would be > perpendicular to the second longest dimension. I've been cutting my > spleens parallel to the longest dimension and perpendicular to the > second longest dimension, because that's where I seem to get the most > information, and it also looks like what I see in my hematopathology > textbook. Is that correct? I know there's probably a standard way > to do it that everybody knows but I'm not a histologist, pathologist > or hematologist and I'm pretty uninformed about the standard clinical > ways to do spleens. I would appreciate any advice, references, etc. > Thanks in advance! > > Sharon > -- > Sharon Cooperman > NIH, NICHD, CBMB 301.435-8417 > Building 18T, room 101 301.402-0078 fax > Bethesda, MD 20892 > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > From Ronnie_Houston <@t> bshsi.com Fri Oct 14 09:53:43 2005 From: Ronnie_Houston <@t> bshsi.com (Houston, Ronnie) Date: Fri Oct 14 09:55:53 2005 Subject: [Histonet] Immunoflouresence Questions Message-ID: <530361BF03351B4CAE5270A05D3037B506B71AAC@bsrexms01.bshsir.com> For those unfamiliar with Mowiol, it is a Polyvinyl Alcohol and can be obtained from Calbiochem. As Tim said, DABCO can be added as an antifading agent (25mg/ml), as can para-phenylene diamine (10mg/ml) and n-propyl gallate (2% w/v); in fact all three agents can be combined in one solution. All these substances are harmful by ingestion, inhalation and skin-absorption. In my opinion, the Alexa dyes from Molecular Probes are much more resistant to photo bleaching than any others on the market, and if memory serves me correctly Molecular Probes do have a commercial mounting medium that contains a reagent (proprietary) to prevent fading. Ronnie Ronnie Houston Director of Anatomic Pathology Bon Secours HealthPartners Laboratories 5801 Bremo Road Richmond, VA 23226 (804) 2877972 (804) 2877906 - fax ronnie_houston@bshsi.com -----Original Message----- From: ana.merino-trigo [mailto:ana.merino-trigo@wanadoo.fr] Sent: Thursday, October 13, 2005 3:49 PM To: Rene J Buesa; Nita Searcy; histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Immunoflouresence Questions Hi, I'm not really an expertise, but in my first postdoc in Australia, I was working in a Cell biology department and we were using Mowiol as mounting medium. We kept the slides in dark at 4?C, and the fluorescence was the same the day after, than the week after, even I was reviewing slides after one month on the fridge, with no a great difference. However, I was working with cell lines, using as FITC, TxRed, Alexa ... antibodies and I don't really now if tissues fades quicker. Attached the protocol used to prepare Mowiol. I do want to take the opportunity to say everyone thank you, I'm learning a lot by reading Histonet disccusions. Great work everyone. Mowiol Mounting Medium pH 8.5 Materials Hopval 5-88 4.8g glycerol 12.0g MilliQ H2O 12.0ml 0.2M Tris-HCl pH 8.5 24.0ml Tris 2.42g Tris base milliQ H2O 100ml pH to 8.5 with 6N HCl Method . Place glycerol in 50ml disposable blue cap tube . Add Hopval and stir thoroughly . Add H2O and leave for 2h at R.T. . Add tris and incubate at ~53?C until the Hopval has dissolved, stirring occassionally (takes ages) . clarify by centrifugation at 3500rpm for 30min and aliquot the supernatant into 1ml in 1.5 ml tubes . store at -20?C, stable at this temp for 12 months . once defrosted, stable at R.T. for one month Thanks Ana ----- Original Message ----- From: Rene J Buesa To: Nita Searcy ; histonet@lists.utsouthwestern.edu Sent: Thursday, October 13, 2005 9:28 PM Subject: Re: [Histonet] Immunoflouresence Questions Nita: You are right, immunofluorescence procedures will fade, and fast; that is the reason behind taking photographs of the results. You may keep the slides in a refrigerator and will be able to register immunofluorescence again, but WEAKER, the next day. How long will this fluorescence will remain visible in refrigerated slides depends on the case, but cannot be predicted in general. When the pathologist that is scheduled to read our procedures is not available, we keep the slides for the next day, but that is all; no permanent solution. There is no media that will retard fading (is something inherent to the FITC conjugated antibody and has nothing to do with the medium. Probably your clinician "heard" something but evidently does not know the procedural details, or he may know something that neither you or I know. Rene J. Nita Searcy wrote: I have a tech that is asking these questions regarding possible "new procedures". We have a new clinician that has "new" ideas. 1. Does storing slides in refrigerator after staining aid in anyway? (I had never seen this before ; always stored in dark @ room temperature). 2. IS there a mounting media that aids in permanence? 3. To my knowledge, these slides are not permanent (no matter what you do) but will inspectors really expect to see them- even in a faded condition? (we take pictures of the kidneys) Thanks _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet -- No virus found in this incoming message. Checked by AVG Anti-Virus. Version: 7.0.344 / Virus Database: 267.11.14/129 - Release Date: 11/10/2005 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ________________________________________________________________________________________________________________________________ ________________________________________________________________________________________________________________________________ The information in this communication is intended to be confidential to the Individual(s) and/or Entity to whom it is addressed. It may contain information of a Privileged and/or Confidential nature, which is subject to Federal and/or State privacy regulations. In the event that you are not the intended recipient or the agent of the intended recipient, do not copy or use the information contained within this communication, or allow it to be read, copied or utilized in any manner, by any other person(s). Should this communication be received in error, please notify the sender immediately either by response e-mail or by phone, and permanently delete the original e-mail, attachment(s), and any copies. From JEllin <@t> yumaregional.org Fri Oct 14 10:50:46 2005 From: JEllin <@t> yumaregional.org (Jesus Ellin) Date: Fri Oct 14 10:51:45 2005 Subject: [Histonet] Grossing Tech Message-ID: What are the going rates for the grossing tech?? Does it vary by specimen size and difficulty??? And how are other facilities categorizing these people?? Do they make more that your regular tech?? Or is as other duties as assigned Jesus Ellin Yuma Regional Medical Center From rocan <@t> mac.com Fri Oct 14 11:35:06 2005 From: rocan <@t> mac.com (Rocan) Date: Fri Oct 14 11:35:44 2005 Subject: [Histonet] Perfusion/Dye In-Reply-To: References: Message-ID: <8F2CEAA0-0269-4BA7-B35F-681E8434F622@mac.com> Hi Katie, You need to incorporate something that does not diffuse outside the vascular walls or gets washed away with the perfusion. We use lectins such as LEA (Licopersicum esculentum agglutinin (Tomato lectin). You must inject before you start the perfusion and allow it to circulate. Dr. Donald McDonald at UCSF has many papers published where they inject such lectin. You may want to see his papers on PubMed. We use about a tenth of the doses he uses and works just fine. All the vessels will be labeled. You can buy LEA labeled with fluorescent molecules, with biotin, HRP, etc. It is not very expensive and the labeling is gorgeous. You can contact me directly if you have any questions. I hope this helps Good luck, Rocio ----- Dr.Rocio Sierra-Honigmann Director Engineered Wound Repair Laboratory Cedars Sinai Research Institute Davis 1091 310-423-1882 Honigmannr@cshs.org On Oct 14, 2005, at 4:41 AM, Crosby, Katherine wrote: > Hi All, > I am looking for suggestions of dyes, fluorescent or otherwise, > that could be incorporated into a standard mouse perfusion procedure. > Thanks in advance, > Katie > > > > > > Katie Crosby > Manager, Immunohistochemistry > Cell Signaling Technology > 166B Cummings Center > Beverly, MA 01915 > > Phone: 978-867-2352 > Fax: 978-867-2402 > E-mail: kcrosby@cellsignal.com > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > From info <@t> instrumedics.com Fri Oct 14 11:57:29 2005 From: info <@t> instrumedics.com (Instrumedics) Date: Fri Oct 14 11:58:41 2005 Subject: [Histonet] Re: Tissue with Fat References: Message-ID: <01b501c5d0e0$75aad9b0$6401a8c0@INSTRUMEDICS22> Arshia, The CryoJaneTape-Transfer system makes it possible to cut fat with remarkable success. The tape adhered to the block face captures the section as you cut the section. You then transfer this intact section to an adhesive coated slide. Please see the details on our web site www.instrumedics.com or contact us with your questions. We have a CD-R that we can send you with a demonstration of the entire CryoJane process from snap-freezing the tissue on the Gentle Jane to cutting the section on to the tape and finally transferring it to the slide. Bernice 800-237-2772 ----- Original Message ----- From: "Arshia Mian" To: Sent: Thursday, October 13, 2005 12:32 PM Subject: [Histonet] Re: Tissue with Fat I have been having some problems with cutting Tissue with Fat on a cryostat (box temp and object temp are at -20C). The slices I cut tend to curl in the middle of the block close to the tissue. I have changed the blade several times, but it still continues to happen even with a new blade. Is there any correlation between the box temp and the object temp that would cause this to happen? Any suggestions? I have also noticed that at times, even though my setting is set to 20 microns, that I get thick and thin cuts. Any suggestions on this as well? I would greatly appreciate it. Thanks. From rjbuesa <@t> yahoo.com Fri Oct 14 12:42:00 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Fri Oct 14 12:42:16 2005 Subject: [Histonet] (no subject) In-Reply-To: <1129290094.434f996e8ed50@webmail.nf.au.dk> Message-ID: <20051014174200.17659.qmail@web61224.mail.yahoo.com> I used the DAKO CD117 (rabbit Ig) at 1:25 with HIER at pH 6.0 in the steamer (20 min in + 20 min out) without any problem (in human tissue with human melanoma control). Are you running a "other than rat" parallel control to be sure that your protocol is working OK? Rene J. b003046@nf.au.dk wrote: Hi, im trying to get the DAKO polyclonal rabbit anti-human c-kit (CD117) to work in rats in paraffin. I have tryed using antigen retrieval in microwave and down to 1:50 dilution. Also I have tryed the C-19 c-kit from Santa Cruz and used both on paraffin and frozen tissue. It just doesnt want to work! Can anybody help? Thanks, Mette K. Hagensen Department of Cardiology B, Research Unit Aarhus University Hospital, Skejby Hospital Brendstrupgaardsvej 100 8200 Aarhus N Denmark Phone: +45 89496237 Fax: +45 89496009 Mail: b003046@nf.au.dk _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From PIXLEYSK <@t> UCMAIL.UC.EDU Fri Oct 14 12:54:16 2005 From: PIXLEYSK <@t> UCMAIL.UC.EDU (Pixley, Sarah (pixleysk)) Date: Fri Oct 14 12:54:27 2005 Subject: [Histonet] RE:immunofluorescence Message-ID: Dear Nita: We currently are fairly happy with Fluoromount G, a water-miscible, permanent-drying mounting medium purchased from Southern Biotech, Cat #0100-01. But Fisher also sells a couple of similar media that do help decrease the fading (Permount and Gel/Mount). They all vary a little, but not by a huge difference. They just slow the photobleaching to a degree. Then we store slides at room temperature in the dark. No advantage to refrigerating and it certainly won't dry quickly in the fridge. You remove excess mounting medium on top and sides of coverslip with water. I have looked at slides after a year or so and if the staining was sufficiently bright to begin with, you can still see some. But I wouldn't want to take pictures then. I don't know one that is used to mount from xylene. But the last time I did that, I just used a standard Depex mounting medium. I think the fading was not too bad with that. Sarah From rjbuesa <@t> yahoo.com Fri Oct 14 12:59:19 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Fri Oct 14 12:59:32 2005 Subject: [Histonet] Dry tissue In-Reply-To: <0E394B648E5284478A6CCB78E5AFDA2718C46B@hpes1.HealthPartners.int> Message-ID: <20051014175919.45291.qmail@web61214.mail.yahoo.com> If nothing has changed in your processing routine then you should look at the biopsies and how they are collected. Once we had a similar problem and it turned out that the biopsies were not fixed immediately they were collected (they dried out before being fixed). Also, are the same fixative as before being used? Have you had any changes in the reagents suppliers? Additionally all our "dryness" problems were eliminated when we start using a mixture of ethanol/isopropanol/mineral oil which allowed us to start dehydration/infiltration steps at a time ratio of 4.8:1 with relation with the initial dehydration. Just a thought! Rene J. "Webb, Dorothy L" wrote: I thought I knew most of the processing answers when hit with problematic tissues to cut, but, lately, I am stymied!! Lately, our smaller biopsies have been very dry and nothing has changed in our processing routine or reagents. I did lessen the times in absolute a little and also let the time in xylene be lessened. I am not seeing the change I thought I would. Can anyone give me any suggestions as to how to process small biopsies in the same processor with large specimens without drying out the small GI biopsies and still getting the proper processing on large speicmens. Thanks ahead of time for responding to this!! ________________________________________ This e-mail and any files transmitted with it are confidential and are intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient or the individual responsible for delivering the e-mail to the intended recipient, please be advised that you have received this e-mail in error and that any use, dissemination, forwarding, printing, or copying of this e-mail is strictly prohibited. If you have received this e-mail in error, please immediately notify the HealthPartners Support Center by telephone at (952) 967-6600. You will be reimbursed for reasonable costs incurred in notifying us. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From cwscouten <@t> myneurolab.com Fri Oct 14 14:16:43 2005 From: cwscouten <@t> myneurolab.com (Charles Scouten) Date: Fri Oct 14 14:17:40 2005 Subject: [Histonet] MOUSE BRAIN FIXATION Message-ID: <5784D843593D874C93E9BADCB87342AB916768@tpiserver03.Coretech-holdings.com> I downloaded and read this article. I am also surprised to read the authors statements that it works so well. I would have some questions and concerns remaining after reading the article. The fixative arrives while the red bloods cells are still present. Don't they crosslink to proteins in the blood vessel linings, and adhere? Does this procedure really wash out red blood cells more cleanly than the traditional procedure? With no prewash? Sufficient to run a clean HRP reaction with red cells catalyzing noise reactions? Where do they go? Since no escape vessel is cut, the excess fluid must accumulate somewhere. There is a clear experience and skill requirement here to press the plunger at the right force and rate. There is a need for an instrument to control the expulsion if this works as well as the authors seem to be saying. I would like to hear from them about this. The authors miss-attribute to Cragg the contention that pressure causes swelling. I can assure them from experience that this is not the case, and will publish on that and other issues shortly . The traditional procedure does indeed produce swelling and distortion, followed by shrinkage, due to fixation of the sodium pump proteins and the inrush of sodium ions. Cragg used a very high pressure, 300 mm Hg, to force isotonic sucrose across the blood brain barrier to get rid of sodium in the extracellular fluid before the fixative arrived, and that avoided the swelling and subsequent shrinkage. I note that the pressures used by the authors in pushing on the syringe plunger is unknown, but great enough to force fluid in against physiological blood pressure. I was impressed that the ventricles in the samples shown were not swollen, much like unfixed, unperfused tissue. How do we know the blood was out? H&E staining works fine in blood filled tissue, and in unfixed tissue. Does GFAP? I guess the EM shows that at least cortex as fixed and preserved. Why is postimmersion fixation unnecesary in this tissue? Both perfusion methods get fixative everywhere fast, but traditional gets progressively better fixations with days in post perfusion fixative. That wouldn't happen with this procedure? If you do try this, I would very much like to hear the outcome. Cordially, Charles W. Scouten, Ph.D. myNeuroLab.com 5918 Evergreen Blvd. St. Louis, MO 63134 Ph: 314 522 0300 x 342 FAX 314 522 0377 cwscouten@myneurolab.com http://www.myneurolab.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Geoff McAuliffe Sent: Thursday, October 13, 2005 1:27 PM To: Favara, Cynthia (NIH/NIAID); Histonet Subject: Re: [Histonet] MOUSE BRAIN FIXATION Years ago my colleagues who worked on mouse bloood would withdraw up to 1 ml of blood from the mouse heart in a similar manner, but hitting the left ventricle does take some practice. I am a little amazed at the low volume of fixative as well, but tilting the animal so that the head is lower than the heart would help. Note that the resolution of the photos is not all that good. Geoff Favara, Cynthia (NIH/NIAID) wrote: >I have this paper and hope to try this next week. Note that the >fixative is picric acid/paraformaldehyde/gluteraldehyde. I would love >to get great fixation with 1ml and without opening the chest cavity. I will let you know! > >c > >Cynthia Favara >NIAID/NIH/RML/LPVD >903 South 4th Street >Hamilton, MT 59840 >406-363-9317 > >-----Original Message----- >From: Adam Perry [mailto:kaleid11@yahoo.com] >Sent: Thursday, October 13, 2005 8:20 AM >To: histonet@lists.utsouthwestern.edu >Subject: [Histonet] MOUSE BRAIN FIXATION > >I downloaded the technical report on this variant perfusion technique. >I'm amazed that you can get good fixation of the brain with 1 ml of >fixative pumped through the heart without clamping off descending >aorta, etc. Has anyone tried this technique (or similar variants), and >what kind of results were obtained? > >I'm planning on trying it in a week or so, because I would love to get >good brain fixation with so little time, fixative and waste >production...but am wary... > >Adam Perry > >Department of Physiology and Biophysics University of Illinois Chicago, >IL 60612 > >There have been several inquiries regarding fixation of mouse brains >recently. > >A short technical report has just been published in BioTechniques: > >Minimally invasive method for murine brain fixation Eichenbaum KD et al >Biotechniques 2005; 39(4): 487-500 > >After simple registration, it can be accessed at >http://www.biotechniques.com > >Ronnie Houston >Director of Anatomic Pathology >Bon Secours HealthPartners Laboratories >5801 Bremo Road >Richmond, VA 23226 >(804) 2877972 >(804) 2877906 - fax >ronnie_houston@bshsi.com > > > > >--------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > -- -- ********************************************** Geoff McAuliffe, Ph.D. Neuroscience and Cell Biology Robert Wood Johnson Medical School 675 Hoes Lane, Piscataway, NJ 08854 voice: (732)-235-4583; fax: -4029 mcauliff@umdnj.edu ********************************************** _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From histo <@t> compbio.com Fri Oct 14 15:45:02 2005 From: histo <@t> compbio.com (Histology) Date: Fri Oct 14 15:47:39 2005 Subject: [Histonet] New Histo Photo Gallery Message-ID: <000201c5d100$2734bce0$4075010a@steveyum> Hello Fellow Histonetters! We have just posted a new histology photo gallery on our website at www.compbio.com. We invite our fellow histonetters to visit our histo photo gallery. Cheers! The histo lab at Comparative Biosciences. From godsgirlnow <@t> msn.com Fri Oct 14 16:04:14 2005 From: godsgirlnow <@t> msn.com (Roxanne Soto) Date: Fri Oct 14 16:04:34 2005 Subject: [Histonet] JOB IN FLORIDA Message-ID: We have opportunities for histotechs that have a florida technologist license----excellent pay. Please contact me directly. Position is for a private Tampa lab... Roxanne From Linda-Boone <@t> ouhsc.edu Fri Oct 14 16:22:31 2005 From: Linda-Boone <@t> ouhsc.edu (Boone, Linda (HSC)) Date: Fri Oct 14 16:22:42 2005 Subject: [Histonet] unsubscribe Message-ID: <80EE593D374DFB42AD9E70B882C4748A5DD4AA@TRIGGER.hsc.net.ou.edu> From clarissabush <@t> sbcglobal.net Fri Oct 14 17:00:08 2005 From: clarissabush <@t> sbcglobal.net (CM Bush) Date: Fri Oct 14 17:00:28 2005 Subject: [Histonet] IHC in Fixed Frozen vs. Processed and Paraffin Embedded Brain Tissue Message-ID: <20051014220008.7707.qmail@web80313.mail.yahoo.com> Hello there Dear Histonet, I am re posting this from a few days ago. The were no responses, so I'm hoping maybe this time. Much thanks for everyone's patience. We are trying to stain human anterior cingulate brain with anti Calretinin. We have taken a sample of said tissue (fixed in 10% NBF); processed and paraffin embedded one portion of the tissue and cut the other portion on a freezing stage microtome (fixed frozen sample cryo protected with 30% sucrose). (The sections for both varieties mounted on slides.) We have been able to see Calretinin staining (at 1:3000) for the processed, embedded tissue but not for the fixed frozen version of the same tissue. For all tissue, antigen retrieval was done using citrate buffer pH6 and microwaving 12 min high and 6 min low, in a pressure cooker. Also, we used TBS buffer with 0.4% Triton X for all sections. Would anyone have any ideas as to what might be going on that the processed/embedded tissue will stain and the fixed frozen version of the same sample does not? Thank you very much, again. Clarissa From ploykasek <@t> phenopath.com Fri Oct 14 17:15:23 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Fri Oct 14 17:16:04 2005 Subject: [Histonet] IHC in Fixed Frozen vs. Processed and Paraffin Embedded Brain Tissue In-Reply-To: <20051014220008.7707.qmail@web80313.mail.yahoo.com> Message-ID: Clarissa, If you are performing the HIER in the microwave pressure cooker on the frozen sections, I think perhaps you are "over-retrieving" those sections. That is a harsh pretreatment to apply to frozen sections. We've had luck on frozen sections that we cut on a cryostat, fix in 10%NBF for 30-60 minutes, retrieve in citrate in a 75 degree water bath for 30 minutes, then perform the IHC. I can't be positive that this will fix your problems, but just something we've tried that works for us. I will say we have had some trouble with calretinin on FS tissue, too. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA > Hello there Dear Histonet, > > I am re posting this from a few days ago. The were no responses, so I'm > hoping maybe this time. Much thanks for everyone's patience. > > We are trying to stain human anterior cingulate brain with anti > Calretinin. We have taken a sample of said tissue (fixed in 10% NBF); > processed and paraffin embedded one portion of the tissue and cut the other > portion on a freezing stage microtome (fixed frozen sample cryo protected > with 30% sucrose). (The sections for both varieties mounted on slides.) > > We have been able to see Calretinin staining (at 1:3000) for the > processed, embedded tissue but not for the fixed frozen version of the same > tissue. For all tissue, antigen retrieval was done using citrate > buffer pH6 and microwaving 12 min high and 6 min low, in a pressure cooker. > Also, we used TBS buffer with 0.4% Triton X for all sections. > > Would anyone have any ideas as to what might be going on that the > processed/embedded tissue will stain and the fixed frozen version of the > same sample does not? > > Thank you very much, again. > > Clarissa > > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail message, including any attachments, is for the sole use of the intended recipients and may contain privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message, or you may call PhenoPath Laboratories, Seattle, WA U.S.A. at (206) 374-9000. From clarissabush <@t> sbcglobal.net Fri Oct 14 20:10:48 2005 From: clarissabush <@t> sbcglobal.net (CM Bush) Date: Fri Oct 14 20:11:09 2005 Subject: [Histonet] IHC in Fixed Frozen vs. Processed and Paraffin Embedded Brain Tissue In-Reply-To: <8C79F22926CF4E0-3D0-9341@MBLK-M03.sysops.aol.com> Message-ID: <20051015011048.81522.qmail@web80302.mail.yahoo.com> Hi Lorie, Thanks for getting back to me. Yes this tissue has been fixing in 10% NBF and cut on a freezing stage microtome (not a cryostat). This tissue has been in 10% NBF for month-years, so the tissue is very well fixed. Right, I guess I should have emphasized this in my post. As well I left out, the reason we are cutting the brain tissue on the freezing stage microtome, is so that we may cut thick (40um) sections, which will flatten out on the slides (thick paraffin sections wrinkle more than will work for what we are doing.) Clarissa bjdewe@aol.com wrote: Why are you doing antigen retrieval on frozen sections? AR is for paraffin tissues that are fixed ... or are your frozens fixed as well? Lorie ******************************************************************* Confidentiality Notice: This e-mail message, including any attachments, is for the sole use of the intended recipient(s)and may contain confidential and privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by reply e-mail and destroy all copies of the original message. ******************************************************************* -----Original Message----- From: CM Bush To: histonet@lists.utsouthwestern.edu Sent: Fri, 14 Oct 2005 15:00:08 -0700 (PDT) Subject: [Histonet] IHC in Fixed Frozen vs. Processed and Paraffin Embedded Brain Tissue .AOLPlainTextBody { margin: 0px; font-family: Tahoma, Verdana, Arial, Sans-Serif; font-size: 12px; color: #000; background-color: #fff; }.AOLPlainTextBody pre { font-size: 9pt;}.AOLInlineAttachment { margin: 10px;}.AOLAttachmentHeader { border-bottom: 2px solid #E9EAEB; background: #F9F9F9;}.AOLAttachmentHeader .Title { font: 11px Tahoma; font-weight: bold; color: #666666; background: #E9EAEB; padding: 3px 0px 1px 10px;}.AOLAttachmentHeader .FieldLabel { font: 11px Tahoma; font-weight: bold; color: #666666; padding: 1px 10px 1px 9px;}.AOLAttachmentHeader .FieldValue { font: 11px Tahoma; color: #333333;} Hello there Dear Histonet, I am re posting this from a few days ago. The were no responses, so I'm hoping maybe this time. Much thanks for everyone's patience.We are trying to stain human anterior cingulate brain with anti Calretinin. We have taken a sample of said tissue (fixed in 10% NBF);processed and paraffin embedded one portion of the tissue and cut the other portion on a freezing stage microtome (fixed frozen sample cryo protected with 30% sucrose). (The sections for both varieties mounted on slides.) We have been able to see Calretinin staining (at 1:3000) for the processed, embedded tissue but not for the fixed frozen version of the same tissue. For all tissue, antigen retrieval was done using citrate buffer pH6 and microwaving 12 min high and 6 min low, in a pressure cooker. Also, we used TBS buffer with 0.4% Triton X for all sections. Would anyone have any ideas as to what might be going on that the processed/embedded tissue will stain and the fixed frozen version of the same sample does not? Thank you very much, again. Clarissa _______________________________________________Histonet mailing listHistonet@lists.utsouthwestern.eduhttp://lists.utsouthwestern.edu/mailman/listinfo/histonet From gintini <@t> buffalo.edu Fri Oct 14 21:42:00 2005 From: gintini <@t> buffalo.edu (Beppe Intini) Date: Fri Oct 14 21:42:13 2005 Subject: [Histonet] cartilage staining Message-ID: Dear Histo-Friends, I hope you can help me again....! Just a quick question: is Safranin-O better than Toluidine blue for cartilage staining? And Why? Thanks Beppe Intini -- Giuseppe Intini Clinical Assistant Professor - Periodontics PhD Candidate - Oral Biology University at Buffalo Buffalo, NY 14214 From PKamalavenkatesh <@t> wockhardtin.com Sat Oct 15 00:17:37 2005 From: PKamalavenkatesh <@t> wockhardtin.com (PKamalavenkatesh@wockhardtin.com) Date: Sat Oct 15 00:17:12 2005 Subject: [Histonet] bone marrow - evaluation- Reply to Cheryl Crowder Message-ID: Dear Crowder, Please find enclosed the details of paper regarding bone marrow collection, evaluation in rats etc.It is also having very good references to bone marrow cytology of small animals especially canines. Also the chapter Bone marrow evaluation in Schalm's Veterinary Haematology, 5 th edn, 23-32, also useful for you hopefully. We are following this methodology for some time and we found this is more useful. Also I suggest you to refer the CD collection from European Society of Toxicopatholgy Collection having a nice presentation from Anne Bolliger. Please get back to me if you have any more queries.Good luck. Dr.P.Kamalavenkatesh M.VSc (Pathology) Wockhardt Research Center New Drug Discovery- Biology Maharastra India. PKamalavenkatesh@wockhardtin.com Cytologic evaluation of bone marrow in rats. Indiactions, methods and normal morphology. Veterinary Clinical Pathology, Vol 33/No 2., 2004 Please Visit our New Corporate Web Site www.wockhardt.com --------------------- Disclaimer ------------------------------------------------------------------------------ Information transmitted by this E-MAIL is proprietary to Wockhardt Ltd. and/ or its Group Companies and/or,its Customers and is intended for use only by the individual or entity to which it is addressed, and may contain information that is privileged, confidential or exempt from disclosure under applicable law. If you are not the intended recipient or it appears that this mail has been forwarded to you without proper authority, you are notified that any use or dissemination of this information in any manner is strictly prohibited. In such cases, please delete this mail from your records. --------------------------------------------------------------------------------------------------------------- From PKamalavenkatesh <@t> wockhardtin.com Sat Oct 15 00:31:32 2005 From: PKamalavenkatesh <@t> wockhardtin.com (PKamalavenkatesh@wockhardtin.com) Date: Sat Oct 15 00:30:49 2005 Subject: [Histonet] Re: spleen sectioning Message-ID: Sharon, I agree with that the sectioning of spleen perpendicular to the long axis is a desired one. I also suggest you to refer the following paper. This contains a standarised methodology for sectioning and trimming of all the organs of mice and rat to nullify the variability among labs and evaluation. Good luck. Revised guides for organ sampling and trimming in rats and mice ? Part 1 A joint publication of the RITA1 and NACAD2 groups Experimental and Toxicologic Pathology Volume 55, Issues 2-3 , 2003, Pages 91-106 Dr.P.Kamalavenkatesh M.VSc (Pathology) Wockhardt Research Center New Drug Discovery- Biology Maharastra India PKamalavenkatesh@wockhardtin.com Please Visit our New Corporate Web Site www.wockhardt.com --------------------- Disclaimer ------------------------------------------------------------------------------ Information transmitted by this E-MAIL is proprietary to Wockhardt Ltd. and/ or its Group Companies and/or,its Customers and is intended for use only by the individual or entity to which it is addressed, and may contain information that is privileged, confidential or exempt from disclosure under applicable law. If you are not the intended recipient or it appears that this mail has been forwarded to you without proper authority, you are notified that any use or dissemination of this information in any manner is strictly prohibited. In such cases, please delete this mail from your records. --------------------------------------------------------------------------------------------------------------- From vanann702 <@t> skmc.gov.ae Sat Oct 15 01:40:06 2005 From: vanann702 <@t> skmc.gov.ae (Anne Van Binsbergen) Date: Sat Oct 15 05:57:00 2005 Subject: [Histonet] mercury Message-ID: <0C44F1AAEE47D54DA4210A60AB206F5E02F6D660@SKMCEMAIL.skmc.gov.ae> Hello Histonetters I am trying to get rid of all Mercury in my lab and have been waging a war with one of the pathologists. He is absolutely determined to retain his 'trusted' B5 for bone marrows and all other odds and ends which he reckons should have a piece in B5 - 'just in case' - he believes that B% is better for IHC than formalin We all agree that the B5 morphology is hard to beat, and that there are various other fixatives which could be used instead For the past 18 mths we have been using Millonigs 10% NB Formalin and are getting stunning results, for both routine H&E's as well as IHC. The key to good IHC is good fixation - timing and fixative What I need is a flurry of responses which include your local legislation regarding heavy metals. I am now trying to do this via OH&S. TIA Annie Anne S van Binsbergen, NDMLT (Haem, Cell Path) Anatomical Pathology Laboratory Sheikh Khalifa Medical City PO BOX 51900 Abu Dhabi, UAE ph: +971 2 6102695 mobile:+971 503162804 email: vanann702@skmc.gov.ae CARPE DIEM! Nobody can make you feel inferior without your permission. DANCE LIKE NO-ONE IS WATCHING! From Myri37 <@t> aol.com Sat Oct 15 06:01:15 2005 From: Myri37 <@t> aol.com (Myri37@aol.com) Date: Sat Oct 15 06:01:32 2005 Subject: [Histonet] need training in histology Message-ID: <1a1.3e840fa3.30823b7b@aol.com> Hi histonetters, First, let me inform you that your website has always been a great help, consulting you has become indispensable. I am a technician in a tissue engineering lab. I end up doing histology in paraffin and resin for the past three years which I enjoyed very much. However?.: Histology in paraffin did not cause me any problem in the other hand the one in resin (methylmetacrylate.spurr and epon) cause me few complications considering the fact that my samples are implants of titanium coated with a layer of hydroxyapatite that?s also covered by a cellular layer rich in collagen fibers. I found problems in the choice of resin , the resin MMA and epon present an important retraction , I make thick sections of 100 um then grind them. But I can't make all of the possible staining for collagen and the tissue newly mineralized I would like to know if you know any information on specialize training in histology as close as possible to my field in another word the inclusion of hard resin of diferent biomaterial covered with tissue. the choice of the appropriate fixative, the choice of dehydratation solvent, the choice of resin the best timing for it, a good interpretation of the microscopic blades and make the difference between coloration artifact and reel element. Could you provide me with title of manuals that will help me answer some of these questions? Or where I can find them? And also if there are courses or training to follow? Thank you very much for your information Myriam B France From rjbuesa <@t> yahoo.com Sat Oct 15 09:02:56 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sat Oct 15 09:03:12 2005 Subject: [Histonet] mercury In-Reply-To: <0C44F1AAEE47D54DA4210A60AB206F5E02F6D660@SKMCEMAIL.skmc.gov.ae> Message-ID: <20051015140256.70183.qmail@web61215.mail.yahoo.com> Any war with a pathologists is easy to wage but difficult to win, mostly depending of the pathologist "power". On the other hand you have to understand their position: they are used to "see", to "identify" things after a certain procedure and feel more "secure" if things don't change from what they used to know, or have been trained into. Having said that, and as you point out, B5 is wonderful for morfology (although it requires "de-mercurization" before the staining) but is not better than neutral buffered formalin for general or special staining, including immunohistochemistry. Even the "humble" NBF is good for anything. Mercury banning is general in the USA. I don't know if you will get the "flurry of responses" that you would like, but you can add mine to the others. Hope it will help! Rene J. Anne Van Binsbergen wrote: Hello Histonetters I am trying to get rid of all Mercury in my lab and have been waging a war with one of the pathologists. He is absolutely determined to retain his 'trusted' B5 for bone marrows and all other odds and ends which he reckons should have a piece in B5 - 'just in case' - he believes that B% is better for IHC than formalin We all agree that the B5 morphology is hard to beat, and that there are various other fixatives which could be used instead For the past 18 mths we have been using Millonigs 10% NB Formalin and are getting stunning results, for both routine H&E's as well as IHC. The key to good IHC is good fixation - timing and fixative What I need is a flurry of responses which include your local legislation regarding heavy metals. I am now trying to do this via OH&S. TIA Annie Anne S van Binsbergen, NDMLT (Haem, Cell Path) Anatomical Pathology Laboratory Sheikh Khalifa Medical City PO BOX 51900 Abu Dhabi, UAE ph: +971 2 6102695 mobile:+971 503162804 email: vanann702@skmc.gov.ae CARPE DIEM! Nobody can make you feel inferior without your permission. DANCE LIKE NO-ONE IS WATCHING! _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From LJApuzzio <@t> msn.com Sat Oct 15 09:44:21 2005 From: LJApuzzio <@t> msn.com (Louis Apuzzio) Date: Sat Oct 15 09:44:38 2005 Subject: [Histonet] Histology Opportunity Message-ID: Hello Histoneters; Would like to find an opportunity in the Northwest for Histology. Would prefer a small private setting. If anyone knows of any availability, I would appreciate in hearing from you. All of you are a "cut" above the rest" Thanks and Peace.. Louis J. Apuzzio From rjbuesa <@t> yahoo.com Sat Oct 15 10:02:34 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sat Oct 15 10:02:46 2005 Subject: [Histonet] cartilage staining In-Reply-To: Message-ID: <20051015150235.24740.qmail@web61219.mail.yahoo.com> Toluidine blue has always worked better for me. Both have a similar chemical composition which for Safranin-O is C20 H19 N4 Cl (MW=351) and for Toluidene Blue is C15 H16 N3 Cl and S (MW=238). Their difference is not in the fact that Toluidine Blue has a smaller mollecular weight, but because Toluidine Blue has Sulfur in its mollecule. Probably this is the reason for a better affinity with the Sulfur present in cartilage. Rene J. Beppe Intini wrote: Dear Histo-Friends, I hope you can help me again....! Just a quick question: is Safranin-O better than Toluidine blue for cartilage staining? And Why? Thanks Beppe Intini -- Giuseppe Intini Clinical Assistant Professor - Periodontics PhD Candidate - Oral Biology University at Buffalo Buffalo, NY 14214 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From Malcolm.McCallum <@t> tamut.edu Sat Oct 15 10:17:03 2005 From: Malcolm.McCallum <@t> tamut.edu (Malcolm McCallum) Date: Sat Oct 15 10:19:02 2005 Subject: [Histonet] histogold Message-ID: I have purchased histogold antibody for identifying vitellogen. Anyone out there used histogold and aware of any of the various problems I might encounter, or precautions I should take so that I do not ruin my samples! Malcolm L. McCallum Assistant Professor Department of Biological Sciences Texas A&M University Texarkana 2600 Robison Rd. Texarkana, TX 75501 O: 1-903-233-3134 H: 1-903-791-3843 Homepage: https://www.eagle.tamut.edu/faculty/mmccallum/index.html ________________________________ From: histonet-bounces@lists.utsouthwestern.edu on behalf of Rene J Buesa Sent: Sat 10/15/2005 9:02 AM To: Anne Van Binsbergen; Histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] mercury Any war with a pathologists is easy to wage but difficult to win, mostly depending of the pathologist "power". On the other hand you have to understand their position: they are used to "see", to "identify" things after a certain procedure and feel more "secure" if things don't change from what they used to know, or have been trained into. Having said that, and as you point out, B5 is wonderful for morfology (although it requires "de-mercurization" before the staining) but is not better than neutral buffered formalin for general or special staining, including immunohistochemistry. Even the "humble" NBF is good for anything. Mercury banning is general in the USA. I don't know if you will get the "flurry of responses" that you would like, but you can add mine to the others. Hope it will help! Rene J. Anne Van Binsbergen wrote: Hello Histonetters I am trying to get rid of all Mercury in my lab and have been waging a war with one of the pathologists. He is absolutely determined to retain his 'trusted' B5 for bone marrows and all other odds and ends which he reckons should have a piece in B5 - 'just in case' - he believes that B% is better for IHC than formalin We all agree that the B5 morphology is hard to beat, and that there are various other fixatives which could be used instead For the past 18 mths we have been using Millonigs 10% NB Formalin and are getting stunning results, for both routine H&E's as well as IHC. The key to good IHC is good fixation - timing and fixative What I need is a flurry of responses which include your local legislation regarding heavy metals. I am now trying to do this via OH&S. TIA Annie Anne S van Binsbergen, NDMLT (Haem, Cell Path) Anatomical Pathology Laboratory Sheikh Khalifa Medical City PO BOX 51900 Abu Dhabi, UAE ph: +971 2 6102695 mobile:+971 503162804 email: vanann702@skmc.gov.ae CARPE DIEM! Nobody can make you feel inferior without your permission. DANCE LIKE NO-ONE IS WATCHING! _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rjbuesa <@t> yahoo.com Sat Oct 15 10:30:33 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sat Oct 15 10:30:46 2005 Subject: [Histonet] IHC in Fixed Frozen vs. Processed and Paraffin Embedded Brain Tissue In-Reply-To: Message-ID: <20051015153033.29429.qmail@web61212.mail.yahoo.com> Clarissa, In our laboratory all the frozen sections destined for IHC were air dried, acetone fixed. The OCT was dissolved in water afterwards, the slides placed in PBS and we did NOT use HIER on them. The objective of HIER, as you perfectly know, is to "undo" the crosslinking consequence of formlin fixation, but if you don't fix in formalin, you don't need HIER. Why don't you try this approach? For us always worked OK. Rene J. Patti Loykasek wrote: Clarissa, If you are performing the HIER in the microwave pressure cooker on the frozen sections, I think perhaps you are "over-retrieving" those sections. That is a harsh pretreatment to apply to frozen sections. We've had luck on frozen sections that we cut on a cryostat, fix in 10%NBF for 30-60 minutes, retrieve in citrate in a 75 degree water bath for 30 minutes, then perform the IHC. I can't be positive that this will fix your problems, but just something we've tried that works for us. I will say we have had some trouble with calretinin on FS tissue, too. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA > Hello there Dear Histonet, > > I am re posting this from a few days ago. The were no responses, so I'm > hoping maybe this time. Much thanks for everyone's patience. > > We are trying to stain human anterior cingulate brain with anti > Calretinin. We have taken a sample of said tissue (fixed in 10% NBF); > processed and paraffin embedded one portion of the tissue and cut the other > portion on a freezing stage microtome (fixed frozen sample cryo protected > with 30% sucrose). (The sections for both varieties mounted on slides.) > > We have been able to see Calretinin staining (at 1:3000) for the > processed, embedded tissue but not for the fixed frozen version of the same > tissue. For all tissue, antigen retrieval was done using citrate > buffer pH6 and microwaving 12 min high and 6 min low, in a pressure cooker. > Also, we used TBS buffer with 0.4% Triton X for all sections. > > Would anyone have any ideas as to what might be going on that the > processed/embedded tissue will stain and the fixed frozen version of the > same sample does not? > > Thank you very much, again. > > Clarissa > > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail message, including any attachments, is for the sole use of the intended recipients and may contain privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message, or you may call PhenoPath Laboratories, Seattle, WA U.S.A. at (206) 374-9000. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From linhines <@t> yahoo.com Sat Oct 15 16:19:25 2005 From: linhines <@t> yahoo.com (Linda Hines) Date: Sat Oct 15 16:20:21 2005 Subject: [Histonet] licensing for histology technician in california Message-ID: <20051015211926.74502.qmail@web33005.mail.mud.yahoo.com> Hi! All I would like to know if there is a license requirement to work in california for histologists. I have been attempting to locate on line, no luck... do any of you that work in california know who to contact? If it is necessary? Thanks in advance. Everyone have a good week-end. Linda Hines HT 602-439-4104 Histo-Techs ON CALL --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From jkiernan <@t> uwo.ca Sat Oct 15 23:14:47 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Sat Oct 15 23:15:07 2005 Subject: [Histonet] cartilage staining References: <20051015150235.24740.qmail@web61219.mail.yahoo.com> Message-ID: <4351D3B7.BA005083@uwo.ca> Dear Beppe (and all others), I think the reason toluidine blue is a more pleasing cartilage stain than safranine O has to do with the shape of the dye molecule. Toluidine blue cations are flat, and can pile up in stacks of two or more, so that each half-sulfate-ester anion in the cartilage matrix can bind more than one toluidine blue cation, resulting in intense staining. The stacking also accounts for a shift in the wavelength of maximum absorption. This results in the pleasing metachromatic red-purple colour of cartilage, mast cell granules and some types of mucus, while cell nuclei (DNA) and basophilic cytoplasms (RNA) are blue. Safranine O (which is always a mixture of isomers, because of the way it's made) has much more bulky cations than toluidine blue. They may form aggregates but they cannot stack up tidily. The dye will (like any other cationic dye) stain cartilage fairly selectively if applied from a solution too acidic to stain DNA (eg at pH below 2). If you don't need a counterstain, that's fine. John Kiernan Anatomy, UWO London, Canada. _______________________ Rene J Buesa wrote: > > Toluidine blue has always worked better for me. Both have a similar chemical composition > which for Safranin-O is C20 H19 N4 Cl (MW=351) and for Toluidene Blue is C15 H16 N3 Cl and S (MW=238). > Their difference is not in the fact that Toluidine Blue has a smaller mollecular weight, but > because Toluidine Blue has Sulfur in its mollecule. Probably this is the reason for > a better affinity with the Sulfur present in cartilage. > Rene J. > > Beppe Intini wrote: > Dear Histo-Friends, > I hope you can help me again....! > Just a quick question: is Safranin-O better than Toluidine blue for > cartilage staining? And Why? > Thanks > Beppe Intini > -- > Giuseppe Intini > Clinical Assistant Professor - Periodontics > PhD Candidate - Oral Biology > University at Buffalo > Buffalo, NY 14214 > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > --------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jkiernan <@t> uwo.ca Sat Oct 15 23:21:47 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Sat Oct 15 23:22:09 2005 Subject: [Histonet] MOUSE BRAIN FIXATION References: <5784D843593D874C93E9BADCB87342AB916768@tpiserver03.Coretech-holdings.com> Message-ID: <4351D55B.4785F5FD@uwo.ca> I like your comments, Charles. I too read the paper by Eichenbaum et al in BioTechniques 39(4): 487-500 (2005) and was surprised that a slow 1 ml intracardiac injection of Somogyi & Takagi's (1982) buffered picrate-formaldehyde-glutaraldehyde cocktail would fix a mouse's brain well enough for EM. Eichenbaum et al describe the technique in great detail, and emphasize that deviations and errors will result in failure. Anecdotally: perfused glutaraldehyde-containing fixatives don't clog blood vessels in the way that formaldehyde-only solutions sometimes do. Several hands-on academic researchers that I knew in bygone days made this assertion. I've never had trouble perfusing any water-based fixative except for bad placement of the needle or cannula, and that happens more often with mice than rats, because of the smaller size. Unduly high perfusion pressure can certainly cause artifacts. I've seen (by EM) huge pericapillary spaces in brain tissue following perfusion done with a syringe. The thumb pressure waa unknown but probably was far to high. Perfusion from a bottle 1 metre above the bench does not induce this artifact. A metre of water is a crude approximation to the pressure in a large artery of a living rodent. Eichenbaum et al injected their fixative through a 27-gauge needle. That's much thinner than the ones used for conventional perfusion, when the emphasis is on getting a substantial flow rate. The thin needle and the long, narrow recommended syringe would make it easier to do the injection slowly. Glutaraldehyde reacts rapidly with proteins, and probably gets rid of the blood-brain barrier in seconds. Geoff McAuliffe is amazed at the small volume of the injected fixative. One ml has to be 20 to 30 percent of the blood volume of a 30 gm mouse. John Kiernan Anatomy, UWO London, Canada -------------------- Charles Scouten wrote: > > I downloaded and read this article. I am also surprised to read the > authors statements that it works so well. I would have some questions > and concerns remaining after reading the article. > > The fixative arrives while the red bloods cells are still present. > Don't they crosslink to proteins in the blood vessel linings, and > adhere? Does this procedure really wash out red blood cells more > cleanly than the traditional procedure? With no prewash? Sufficient to > run a clean HRP reaction with red cells catalyzing noise reactions? > Where do they go? Since no escape vessel is cut, the excess fluid must > accumulate somewhere. There is a clear experience and skill requirement > here to press the plunger at the right force and rate. There is a need > for an instrument to control the expulsion if this works as well as the > authors seem to be saying. I would like to hear from them about this. > > The authors miss-attribute to Cragg the contention that pressure causes > swelling. I can assure them from experience that this is not the case, > and will publish on that and other issues shortly . The traditional > procedure does indeed produce swelling and distortion, followed by > shrinkage, due to fixation of the sodium pump proteins and the inrush of > sodium ions. Cragg used a very high pressure, 300 mm Hg, to force > isotonic sucrose across the blood brain barrier to get rid of sodium in > the extracellular fluid before the fixative arrived, and that avoided > the swelling and subsequent shrinkage. I note that the pressures used > by the authors in pushing on the syringe plunger is unknown, but great > enough to force fluid in against physiological blood pressure. > > I was impressed that the ventricles in the samples shown were not > swollen, much like unfixed, unperfused tissue. How do we know the blood > was out? H&E staining works fine in blood filled tissue, and in unfixed > tissue. Does GFAP? I guess the EM shows that at least cortex as fixed > and preserved. Why is postimmersion fixation unnecesary in this tissue? > Both perfusion methods get fixative everywhere fast, but traditional > gets progressively better fixations with days in post perfusion > fixative. That wouldn't happen with this procedure? > > If you do try this, I would very much like to hear the outcome. > > Cordially, > Charles W. Scouten, Ph.D. > myNeuroLab.com > 5918 Evergreen Blvd. > St. Louis, MO 63134 > Ph: 314 522 0300 x 342 > FAX 314 522 0377 > cwscouten@myneurolab.com > http://www.myneurolab.com > > > -----Original Message----- > From: histonet-bounces@lists.utsouthwestern.edu > [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Geoff > McAuliffe > Sent: Thursday, October 13, 2005 1:27 PM > To: Favara, Cynthia (NIH/NIAID); Histonet > Subject: Re: [Histonet] MOUSE BRAIN FIXATION > > Years ago my colleagues who worked on mouse bloood would withdraw up to > 1 ml of blood from the mouse heart in a similar manner, but hitting the > left ventricle does take some practice. > I am a little amazed at the low volume of fixative as well, but tilting > the animal so that the head is lower than the heart would help. > Note that the resolution of the photos is not all that good. > > Geoff > > Favara, Cynthia (NIH/NIAID) wrote: > > >I have this paper and hope to try this next week. Note that the > >fixative is picric acid/paraformaldehyde/gluteraldehyde. I would love > >to get great fixation with 1ml and without opening the chest cavity. I > will let you know! > > > >c > > > >Cynthia Favara > >NIAID/NIH/RML/LPVD > >903 South 4th Street > >Hamilton, MT 59840 > >406-363-9317 > > > >-----Original Message----- > >From: Adam Perry [mailto:kaleid11@yahoo.com] > >Sent: Thursday, October 13, 2005 8:20 AM > >To: histonet@lists.utsouthwestern.edu > >Subject: [Histonet] MOUSE BRAIN FIXATION > > > >I downloaded the technical report on this variant perfusion technique. > > >I'm amazed that you can get good fixation of the brain with 1 ml of > >fixative pumped through the heart without clamping off descending > >aorta, etc. Has anyone tried this technique (or similar variants), and > > >what kind of results were obtained? > > > >I'm planning on trying it in a week or so, because I would love to get > >good brain fixation with so little time, fixative and waste > >production...but am wary... > > > >Adam Perry > > > >Department of Physiology and Biophysics University of Illinois Chicago, > > >IL 60612 > > > >There have been several inquiries regarding fixation of mouse brains > >recently. > > > >A short technical report has just been published in BioTechniques: > > > >Minimally invasive method for murine brain fixation Eichenbaum KD et al > > >Biotechniques 2005; 39(4): 487-500 > > > >After simple registration, it can be accessed at > >http://www.biotechniques.com > > > >Ronnie Houston > >Director of Anatomic Pathology > >Bon Secours HealthPartners Laboratories > >5801 Bremo Road > >Richmond, VA 23226 > >(804) 2877972 > >(804) 2877906 - fax > >ronnie_houston@bshsi.com > > > > > > > > > >--------------------------------- > > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. > >_______________________________________________ > >Histonet mailing list > >Histonet@lists.utsouthwestern.edu > >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > >_______________________________________________ > >Histonet mailing list > >Histonet@lists.utsouthwestern.edu > >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > > > > > > -- > -- > ********************************************** > Geoff McAuliffe, Ph.D. > Neuroscience and Cell Biology > Robert Wood Johnson Medical School > 675 Hoes Lane, Piscataway, NJ 08854 > voice: (732)-235-4583; fax: -4029 > mcauliff@umdnj.edu > ********************************************** > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From vanann702 <@t> skmc.gov.ae Sun Oct 16 02:21:25 2005 From: vanann702 <@t> skmc.gov.ae (Anne Van Binsbergen) Date: Sun Oct 16 02:20:32 2005 Subject: [Histonet] Processing Eyes Message-ID: <0C44F1AAEE47D54DA4210A60AB206F5E02F6D666@SKMCEMAIL.skmc.gov.ae> Hi there Denise Although I am by no means consider myself an 'eye expert' I have had my methods for fixation and processing whole human eyes (and pig) published by 'Microscopy Today' as well as on the histonet over the years. What do you need to know? I will try to help as best I can Cheers Annie Anne S van Binsbergen, NDMLT (Haem, Cell Path) Anatomical Pathology Laboratory Sheikh Khalifa Medical City PO BOX 51900 Abu Dhabi, UAE ph: +971 2 6102695 mobile:+971 503162804 email: vanann702@skmc.gov.ae CARPE DIEM! Nobody can make you feel inferior without your permission. DANCE LIKE NO-ONE IS WATCHING! -----Original Message----- From: Denise Piontek [mailto:dbpiontek@hotmail.com] Sent: Sunday, September 11, 2005 8:40 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Processing Eyes I know I have seen an eye expert on the histonet a few times. I am looking for information on eye fixation and processing for multiple species. Opinions seem to be quite varied. Thanks for your help, Denise Bland-Piontek, HTL(ASCP)CTBNS(AATB) Histology Supervisor TOXiKON _________________________________________________________________ [1]Make FREE PC-to-PC calls with MSN Messenger. Get it now! References 1. http://g.msn.com/8HMAENUS/2749??PS=47575 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From CrochiereSteve <@t> aol.com Sun Oct 16 09:01:36 2005 From: CrochiereSteve <@t> aol.com (CrochiereSteve@aol.com) Date: Sun Oct 16 09:02:36 2005 Subject: [Histonet] micro chatter in prostate bx's Message-ID: <84.4feac465.3083b740@aol.com> I know this has been discussed before, but, does anyone have any suggestions to reduce the amount of microchatter in the glands of prostate needle bx's? I have been having a recurring problem with this (usually with the same tech) and would like to make suggestions to this tech (other than "Don't cut bx's). I've tried soaking the blocks and cutting slower, but this doesn't seem to be solving the issue. Is there a different fixative other than NBF that we could use in the collection kits that may help? Any and all suggestions are welcome. Thanx, Steven M. Crochiere, HT(ASCP) Histology Supervisor LifePath Partners @ Mercy Medical Center Springfield, MA 01104 From rjbuesa <@t> yahoo.com Sun Oct 16 09:43:04 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sun Oct 16 09:43:22 2005 Subject: [Histonet] micro chatter in prostate bx's In-Reply-To: <84.4feac465.3083b740@aol.com> Message-ID: <20051016144304.38402.qmail@web61222.mail.yahoo.com> Usually micro chatter in prostate biopsies (as well as in any other small biopsy, like gastric or colonic) are a result of a combination of factors: 1-they may start at the collection site if the biopsy is let to dry before it is placed in the fixative; 2- NBF is good, but not other alcohol containing fixatives that will start dehydration along with fixation. If the fixative is changed to one containing alcohol, then the processing protocol has to be change to reduce dehydration times; 3-If the biopsies are run along with other general tissues they may run the risk of being overdehydrated; 4-overdehydration will be lessen if infiltration steps are adequate (we overcame these problems when we began infiltrating with mineral oil instead of xylene); 5- another point where the problem can be increased is while casting the blocks if the paraffin does not solidify quickly and completely around the biopsy. 6- cutting too fast can also produce the chatter, as well as an inadequate blade angle. All of the above will be invalidated if you are having problems with only one histotech. If this is the case then the problem should be in the cutting technique. Some "soaking" of the block after it is faced down may help. You should compare the techniques of all your histotechs and try that the one with problems adopts the correct cutting procedure. Also remember that sometimes there are people that just "are not born to cut" (I have had at least 2 of them); this aspect of our art is very tricky (as you know). Hope this will help (although I am sure you knew of all this)! Rene J. CrochiereSteve@aol.com wrote: I know this has been discussed before, but, does anyone have any suggestions to reduce the amount of microchatter in the glands of prostate needle bx's? I have been having a recurring problem with this (usually with the same tech) and would like to make suggestions to this tech (other than "Don't cut bx's). I've tried soaking the blocks and cutting slower, but this doesn't seem to be solving the issue. Is there a different fixative other than NBF that we could use in the collection kits that may help? Any and all suggestions are welcome. Thanx, Steven M. Crochiere, HT(ASCP) Histology Supervisor LifePath Partners @ Mercy Medical Center Springfield, MA 01104 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Sun Oct 16 10:07:51 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sun Oct 16 10:38:49 2005 Subject: [Histonet] cartilage staining In-Reply-To: <4351D3B7.BA005083@uwo.ca> Message-ID: <20051016150751.40111.qmail@web61213.mail.yahoo.com> Dear Beppe (and other): According with Bancroft & Stevens (1977) when discussing the Standard Toluidine Blue (S.T.B.) method by Wolman (1971) they attribute the cartilage methachromatic staining to "probably" an electrochemical bonding that, according with Cooper (1974) is minimised if the staining solution is saturated with sodium chloride. The electrochemical bonding has to occur between the sulfur in the toluidin blue molecule and the chondroitin sulfates molecules found in cartilage. Just an additional detail! Rene J. John Kiernan wrote: Dear Beppe (and all others), I think the reason toluidine blue is a more pleasing cartilage stain than safranine O has to do with the shape of the dye molecule. Toluidine blue cations are flat, and can pile up in stacks of two or more, so that each half-sulfate-ester anion in the cartilage matrix can bind more than one toluidine blue cation, resulting in intense staining. The stacking also accounts for a shift in the wavelength of maximum absorption. This results in the pleasing metachromatic red-purple colour of cartilage, mast cell granules and some types of mucus, while cell nuclei (DNA) and basophilic cytoplasms (RNA) are blue. Safranine O (which is always a mixture of isomers, because of the way it's made) has much more bulky cations than toluidine blue. They may form aggregates but they cannot stack up tidily. The dye will (like any other cationic dye) stain cartilage fairly selectively if applied from a solution too acidic to stain DNA (eg at pH below 2). If you don't need a counterstain, that's fine. John Kiernan Anatomy, UWO London, Canada. _______________________ Rene J Buesa wrote: > > Toluidine blue has always worked better for me. Both have a similar chemical composition > which for Safranin-O is C20 H19 N4 Cl (MW=351) and for Toluidene Blue is C15 H16 N3 Cl and S (MW=238). > Their difference is not in the fact that Toluidine Blue has a smaller mollecular weight, but > because Toluidine Blue has Sulfur in its mollecule. Probably this is the reason for > a better affinity with the Sulfur present in cartilage. > Rene J. > > Beppe Intini wrote: > Dear Histo-Friends, > I hope you can help me again....! > Just a quick question: is Safranin-O better than Toluidine blue for > cartilage staining? And Why? > Thanks > Beppe Intini > -- > Giuseppe Intini > Clinical Assistant Professor - Periodontics > PhD Candidate - Oral Biology > University at Buffalo > Buffalo, NY 14214 > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > --------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Sun Oct 16 10:55:15 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sun Oct 16 10:55:24 2005 Subject: [Histonet] TOT: second update summary. Message-ID: <20051016155515.18282.qmail@web61218.mail.yahoo.com> To all Histonet subscribers: Between 4 and 15 October 2005 I have received the following TOTs: 7-heat the water for the water bath in a microwaves oven to get a faster start to cutting; 8-add a drop of detergent in the water bath to help fatty/poorly processed tissue stay together; 9-if block isn't ribboning well, draw a razor blade/fingernail accross the top and bottom edges to "even them" out; 10-use dryer sheets (like Bounce) to wipe down cryostat of microtome if you are having problems with static electricity; 11-place a piece of pipe insulation on the handle of manual microtomes to avoid repetitive injuries from holding the hand closed too tightly; 12-place heavy pieces of equipment on turntales ("lazy Susans") to reach the back of the instrument easier. 13- use plastic slide mailers to stain small number of slide (saves reagents); 14-use your PC printer to make legible labels of anything; 15-for sections that fall from slides, place slides at 70 Celsius for 30 minutes and immediately after in the freezer; repeat this 2 steps two or three times; 16-air bubbles can be removed from the underside of a floating section by approaching the free end of a tightly rolled piece of paraffin (waste from rough of the block face) held with with a thin forceps to the air bubble. The bubble will be incorporated to the air trapped in the paraffin roll; 17-when doing IHC in animal tissue, test all antibodies in human tissue first; 18-for IHC always prepare reagents fresh every day and discard excess; 19- always prepare 20% more antibody than you need in case you have to repeat or add slides; 20-always read carefully and follow antibody manufacturers' instructions on storage conditions; 21- always purchase antibodies from reputable suppliers; 22-after the blocks have been faced down, place them in a large block of ice before taking the final sections. This second update contains 2.5 times more TOTs than the first but not quite what I was expecting. There are many more TOTs amongst the Histonet subscribers. It will just take a moment to e-mail them, and for sure they will be of benefit for somebody. There is not such a thing as a "foolish" TOT; remember that what for somebody is just "irrelevant routine" could be a treasure for another person. Rene J. Buesa --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Sun Oct 16 13:05:29 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sun Oct 16 13:05:38 2005 Subject: [Histonet] TOTs dealing with TOE NAILS. Message-ID: <20051016180530.87561.qmail@web61214.mail.yahoo.com> To all the Histonet subscribers: Toe Nails has been a subject of frequent discussions in Histonet, and I have been able to find 35 TOTs dealing with this subject since May/99 in Histonet Archieves. Before begin editing these TOTs I want to present them all to you so you can add any one not included, in a way that the final edition will be more comprehensive. All the TOTs found and those that you can send me will be credited to the author, along with the date they were posted. A-Pre-processing ACID hydrolisis (3 TOTs): -use 2% aq. phenol solution overnight; -use 4% alcoholic phenol solution; -use concentrated sulfuric acid for 2-3 days; B-Pre-processing BASIC hydrolisis (11 TOTs): -macerate in 40% aq. sodium hydroxide for 3 hours; -use 10% aq. sodium hydroxide for 30 minutes with constant agitation; -use 5% aq. sol. of sodium hydroxide; -use 15% ammonia water; -use calcium hydroxide; -use depilatory NAIR for a few days until toe nail is pliable; -use NAIR for at least 24 hours; -use softener MOLLIFLEX avoiding tissue to swell; -use diluted solution of fabric conditioner; -soak after grossing during several hours in "chitin" treatment; -use warm soapy water; C-Depilatories and softeners (3 TOTs): -use MOLLIFEX after processing (never before); -depilatories NAIR and NEET will cause weaker staining; -NAIR does not work really well; D-Blocks treatment (12 TOTs): -ice the blocks really well after trimming; -soak blocks overnight in ice water; -soak blocks in warm soapy water; -use 15% ammonia water; -treat trimmed surface with MOLLIFEX; -treat block with "chitin softening solution"; -soak the trimmed block with Downy fabric softener for about 5 min.; -use NAIR on the trimmed block; -use NAIR for 5-10 min., but always less than 15 min.; -place blocks over a frozen block of 0.2% fabric softener; -soak blocks in soapy water (a few drops of Dawn dish washer in 10-20 mL of water; -use MOLLIFEX on trimmed block surface; E-Sections of toe nails (6 TOTs): -add wood glue to the water bath; -coat slides with a 5% aq. sol. of commercial wood glue; -always use + charged slides; -use + charged slides with smeared Elmer's glue; -use a coat of 5% aq. sol. of yellow wood glue; -heat the water bath to 70 Celsius in a way that the toe nail section can float freely after the paraffin has melted at that temperature; Some of the TOTs are contradictory so in the dinal edition I will deal with the fundamental principle of each group. This and any other summary has to be the result of a colective effort from all of us to those that have less experience of less access to information. I hope to receive your contributions to the summary of the TOTs dealing with the "horny" aspect of TOE NAILS. Rene J. Buesa --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From MTitford <@t> aol.com Sun Oct 16 14:47:24 2005 From: MTitford <@t> aol.com (MTitford@aol.com) Date: Sun Oct 16 14:47:40 2005 Subject: [Histonet] Surprise CAP inspections Message-ID: <1f0.4672bd99.3084084c@aol.com> Reviewing my Histonet mail for the last few days I came across Susan's email asking for advice about CAP inspections. I don't know if anyone mentioned that some time in the future they are about to become surprise inspections. That may be quite a trip. Every laboratory I have worked in traditionally cleaned up the lab when the CAP was coming, reviewed all the manuals, checked the fridge for outdated chemicals, etc. We also set aside a conference room for the inspectors to camp out in, arranged lunch and made appointments for them with the chief of medical staff, hospital administrator, etc. Now we all know that our labs should be tidy all the time, everything in date in the fridge, and we should work like the CAP was coming every day, but there may be some surprises for the CAP inspectors in the future. Key people may be off (Chief pathologist/Hospital administrator/histology supervisor). there may be no lunch for them, no where for them to park etc. In fact they may find no one is ready for them. The state inspectors for medical laboratories in Alabama used to arrive unannounced and it was a trip. Maybe like the Titanic! Any ideas anyone?! Mike Titford USA Pathology Mobile AL USA From rjbuesa <@t> yahoo.com Sun Oct 16 15:20:13 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sun Oct 16 15:20:23 2005 Subject: [Histonet] Surprise CAP inspections In-Reply-To: <1f0.4672bd99.3084084c@aol.com> Message-ID: <20051016202013.84242.qmail@web61216.mail.yahoo.com> In Florida we used to have "surprise" inspections by the State. We were caught "off guard" once and it was not very nice, but not that bad either. At that point I decided to develop a routine that allowed me to "self inspect" weekly, one area at a time. That is what Supervisors are for. I tried to keep things in order. It turned out that after that "surprise visit" the State inspections stopped altogether and we were left with the CAP only that always were announced visits. In short: develop a self inspection routine that will allow you to "feel safe"; besides running the lab correctly is also the goal. Nevermind if they don't have lunch, or parking space; that is the price they will have to pay for having an unannounced inspection. Rene J. MTitford@aol.com wrote: Reviewing my Histonet mail for the last few days I came across Susan's email asking for advice about CAP inspections. I don't know if anyone mentioned that some time in the future they are about to become surprise inspections. That may be quite a trip. Every laboratory I have worked in traditionally cleaned up the lab when the CAP was coming, reviewed all the manuals, checked the fridge for outdated chemicals, etc. We also set aside a conference room for the inspectors to camp out in, arranged lunch and made appointments for them with the chief of medical staff, hospital administrator, etc. Now we all know that our labs should be tidy all the time, everything in date in the fridge, and we should work like the CAP was coming every day, but there may be some surprises for the CAP inspectors in the future. Key people may be off (Chief pathologist/Hospital administrator/histology supervisor). there may be no lunch for them, no where for them to park etc. In fact they may find no one is ready for them. The state inspectors for medical laboratories in Alabama used to arrive unannounced and it was a trip. Maybe like the Titanic! Any ideas anyone?! Mike Titford USA Pathology Mobile AL USA _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From louise.renton <@t> gmail.com Mon Oct 17 02:08:37 2005 From: louise.renton <@t> gmail.com (louise renton) Date: Mon Oct 17 02:08:51 2005 Subject: [Histonet] micro chatter in prostate bx's In-Reply-To: <84.4feac465.3083b740@aol.com> References: <84.4feac465.3083b740@aol.com> Message-ID: This might be stating the obvious but........ If this is a problem with one tech's cutting - does that one person cut on the same microtome each time? If they do, then that may indicate "mechanical" rather than personal error. 1. If you are using tissue tek type embedding moulds and cassettes check that the wax is filled up sufficiently in the cassette to provide good stability. 2. Check that the microtome block clamp is holding the block tightly, and that blades, knives etc are also without "play" 3. Depending on the age of the machine it might need a good service to replace worn parts. regards On 10/16/05, CrochiereSteve@aol.com wrote: > I know this has been discussed before, but, does anyone have any suggestions > to reduce the amount of microchatter in the glands of prostate needle bx's? I > have been having a recurring problem with this (usually with the same tech) > and would like to make suggestions to this tech (other than "Don't cut bx's). > I've tried soaking the blocks and cutting slower, but this doesn't seem to be > solving the issue. Is there a different fixative other than NBF that we could > use in the collection kits that may help? Any and all suggestions are welcome. > Thanx, > > Steven M. Crochiere, HT(ASCP) > Histology Supervisor > LifePath Partners @ Mercy Medical Center > Springfield, MA 01104 > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > -- Louise Renton Bone Research Unit University of the Witwatersrand Johannesburg South Africa "....onwards through the fog!" From Jacqueline.Malam <@t> rli.mbht.nhs.uk Mon Oct 17 03:14:59 2005 From: Jacqueline.Malam <@t> rli.mbht.nhs.uk (Malam Jacqueline) Date: Mon Oct 17 03:18:00 2005 Subject: [Histonet] Re: microchatter Message-ID: I have noticed that, if blocks with glandular components are trimmed then put on an ice tray that has not had time to thaw and produce a film of water on the surface (either naturally or by pouring some water onto the tray), microchatters are more likely to form - ie, the blocks are too cold. Hope this helps Jacqui Malam Lancaster DISCLAIMER: This e-mail is confidential and privileged. If you are not the intended recipient please accept our apologies; please do not disclose, copy or distribute information in this e-mail or take any action in reliance on its contents: to do so is strictly prohibited and may be unlawful. Please inform postmaster@rli.mbht.nhs.uk that this message has gone astray before deleting it. Comments or opinions expressed in this email are those of their respective contributors only. The views expressed do not represent the views of the Trust, its management or employees. Morecambe Bay Hospitals NHS Trust is not responsible and disclaims any and all liability for the content of comments written within.Thank you for your co-operation. From pex0220 <@t> yahoo.com.cn Mon Oct 17 07:16:22 2005 From: pex0220 <@t> yahoo.com.cn (pex) Date: Mon Oct 17 07:16:43 2005 Subject: [Histonet] Immunohistochemistry questions Message-ID: <20051017121622.84068.qmail@web15507.mail.cnb.yahoo.com> Dear all, I am confused about immunohistochemistry. I have had some nice results in immunohistochemistry of paraffin-embedded bone and cartilage sections with alkaline phosphatase antibody. But there are not some good results in bone sections if I do immunofluorescence with the same antibody, I do not know the reasons. Maybe due to too much autofluorescence, but I am not sure. Any suggestions will be helpful. Thanks a lot. Guofeng --------------------------------- ÑÅ»¢Ãâ·ÑGÓÊÏ䣭ÖйúµÚÒ»¾øÎÞÀ¬»øÓʼþɧÈų¬´óÓÊÏä ÑÅ»¢ÖúÊÖ¡§DËÑË÷¡¢É±¶¾¡¢·ÀɧÈÅ From LJApuzzio <@t> msn.com Mon Oct 17 07:27:33 2005 From: LJApuzzio <@t> msn.com (Louis Apuzzio) Date: Mon Oct 17 07:27:50 2005 Subject: [Histonet] Histology Opportunity Message-ID: Hello Histonetters; Is anyone aware of any current openings in Histology ? I am currently in Oregon and am looking for a more solid work environment. Would appreciate your help !! Thanks and have a good day !! Louis J. Apuzzio From anh2006 <@t> med.cornell.edu Mon Oct 17 07:27:46 2005 From: anh2006 <@t> med.cornell.edu (Andrea Hooper) Date: Mon Oct 17 07:28:02 2005 Subject: [Histonet] Immunohistochemistry questions Message-ID: <74cb11def488.43536082@med.cornell.edu> If you look at your sections with no primary and no secondary you should be able to determine if it is autofluorescence or not. It is my experience that paraffin embedded bone is highly autofluorescent and needs to be blocked prior to staining. If you provide more details of your protocol perhaps we can help. From Inga.Hansson <@t> neuro.uu.se Mon Oct 17 07:43:53 2005 From: Inga.Hansson <@t> neuro.uu.se (Inga Hansson) Date: Mon Oct 17 07:35:38 2005 Subject: [Histonet] Anti-galanin Message-ID: <531f321a3d1caa9bf0f89be7abc1ef54@neuro.uu.se> Hi there Is anyone familiar with an antibody against galanin that works on mouse paraffin sections?? Thanks in advance! Inga From sjchtascp <@t> yahoo.com Mon Oct 17 07:51:52 2005 From: sjchtascp <@t> yahoo.com (Steven Coakley) Date: Mon Oct 17 07:52:01 2005 Subject: [Histonet] Antigen retrieval Message-ID: <20051017125152.79060.qmail@web90207.mail.scd.yahoo.com> Has anyone had problems with unwanted specific nuclear staining after performing heat AR especially from the microwave. I've been attempting to stain paraffin embedded rat liver in which a 24hour and 48hour BDL with Brdu injection 1 hour prior to harvest. With any form of AR, even limited 80C I get 75-100% nuclear staining. With no AR I get nothing, not even the bile ducts. Does AR tend to effect nuclear staining with some antibodies. I'm stumped. Steve --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rschoon <@t> email.unc.edu Mon Oct 17 08:18:29 2005 From: rschoon <@t> email.unc.edu (rschoon) Date: Mon Oct 17 08:19:15 2005 Subject: [Histonet] Antigen retrieval In-Reply-To: <20051017125152.79060.qmail@web90207.mail.scd.yahoo.com> References: <20051017125152.79060.qmail@web90207.mail.scd.yahoo.com> Message-ID: <4353A4A5.20803@email.unc.edu> Steve, The following procedure has worked in my lab for years and has also been published as it has been used in the publication of several articals and reports. If the directions are followed faithfully you will have excellent results without nonspecific background. Regards, Robert Schoonhoven *_BrdU Immunohistochemistry for use with Dako Envision^(TM) HRP Kit_* *Antibody: anti BrdU* Figure A: *Rat Kidney, 200x.* Clone: Bu20a Supplier: Dako Catalog No.: M 0744 Ig Species: Mouse IHC Method: Formalin - Paraffin, Frozen, Plastic GMA & MMA Pretreatment: see method (below, 1-5) IHC Handling: Manual Isotype: IgG1 Procedure Species: all mammals IgG Concentration: 1mg/ml *_ _* *_Specificity: Proliferating Cells of all Mammals Treated with Bromodeoxyuridine_* * * *_Staining Pattern: Nuclear _* *_ _* *_Procedure:_* *_ _* 1) Hydrate slides to ddH_2 O as per SOP. 2) Hydrolyze with 4N HCl at 37^o C for 20 minutes. 3) Rinse with ddH_2 O for 1 minute at RT. 4) Transfer slides to a staining dish filled with ddH_2 O (kept at 37^o C) for 5 minutes. 5) Incubate in pepsin solution (Dako) for 15 minutes at 37^o C. *_All of the remaining steps are carried out at room temperature._* 6) Rinse 2X with ddH_2 O, 1 minute each rinse. 7) Rinse 2X with PBSt, 3 minutes each rinse 8) Peroxidase Blocking Reagent (Dako), 5 minute incubation. 9) Repeat step #7. 10) Primary antibody (anti-BrdU), incubate 10 minutes at a 1:200 dilution. [5 ?L antibody to 995 ?L 1% BSA in PBS] 11) Repeat step #7. BrdU Page 1 of 2 12) Polymer labeled secondary antibody (Dako Envision HRP), incubate for 10 minutes. 13) Repeat step #7. 14) Incubate in working Dako DAB solution for 8 minutes (1 drop DAB Chromogen per 1 ml substrate buffer). 15) Rinse well with ddH_2 O. 16) DAB enhancer solution (2.5% CoCl_2 , 2.0% NiSO_4 (NH_4 )_2 SO_4 in ddH_2 O) for 8 minutes 17) Repeat step #15. 18) Richard Allan Hematoxylin 1 for 25 seconds, rinse with tap water until there is no blue color in the water. 19) Let sit in tap water for 5 minutes. 20) dehydrate and coverslip in accordance with SOP's. /_Method developed by: Robert Schoonhoven_/ /_ _/ /_S.O.P. Effective January 13, 2003_/ BrdU Page 2 of 2__ Steven Coakley wrote: >Has anyone had problems with unwanted specific nuclear staining after performing heat AR especially from the microwave. I've been attempting to stain paraffin embedded rat liver in which a 24hour and 48hour BDL with Brdu injection 1 hour prior to harvest. With any form of AR, even limited 80C I get 75-100% nuclear staining. With no AR I get nothing, not even the bile ducts. Does AR tend to effect nuclear staining with some antibodies. I'm stumped. > >Steve > > >--------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > From sluhisto <@t> yahoo.com Mon Oct 17 09:04:43 2005 From: sluhisto <@t> yahoo.com (Histology SLU) Date: Mon Oct 17 09:04:53 2005 Subject: [Histonet] Block cutting protocols Message-ID: <20051017140443.66426.qmail@web51010.mail.yahoo.com> Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From sluhisto <@t> yahoo.com Mon Oct 17 09:07:40 2005 From: sluhisto <@t> yahoo.com (Histology SLU) Date: Mon Oct 17 09:07:51 2005 Subject: [Histonet] Histology Opportunity In-Reply-To: Message-ID: <20051017140740.67119.qmail@web51010.mail.yahoo.com> Louis: We do not have anything right now here in St. Louis, Missouri, but there is potential for the future. If you would be interested in sending me your CV, I would be happy to hold it until such time as we might have an open position. You can forward to this email address or fax to 314-977-8740. Have a super week. Susan Louis Apuzzio wrote: Hello Histonetters; Is anyone aware of any current openings in Histology ? I am currently in Oregon and am looking for a more solid work environment. Would appreciate your help !! Thanks and have a good day !! Louis J. Apuzzio _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Mon Oct 17 09:25:22 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Mon Oct 17 09:25:30 2005 Subject: [Histonet] Block cutting protocols In-Reply-To: <20051017140443.66426.qmail@web51010.mail.yahoo.com> Message-ID: <20051017142522.58135.qmail@web61216.mail.yahoo.com> That practice is absolutely forbiden. It can lead to one of the most dangerous mistakes in pathology, which is having the sections from one patient assigned to another, with all the horror diagnostic scenarios, and their consequences, that you can imagine. This was done twice "in my watch" and the histotech got two written counsellings that lead to a termination later on (for another reason). In both cases there were no patient consequences because the type of tissue was not the "right" one as described in the sample, but it was a "close call" and could have been very very bad. This is an absolute "no, no" and due to that experience, we had a meeting, and all new employees were aware of that mistake, and this "no, no" was incorporated to our SOP. Rene J. Histology SLU wrote: Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rschoon <@t> email.unc.edu Mon Oct 17 09:42:36 2005 From: rschoon <@t> email.unc.edu (rschoon) Date: Mon Oct 17 09:43:09 2005 Subject: [Histonet] Major Oops Re:BrdU Antigen retrieval In-Reply-To: <4353A4A5.20803@email.unc.edu> References: <20051017125152.79060.qmail@web90207.mail.scd.yahoo.com> <4353A4A5.20803@email.unc.edu> Message-ID: <4353B85C.7000402@email.unc.edu> My aplogies to the list. Tried to cut and paste so that everyone would get it but that didn't work out so I sent it to Steve as an attachment. Regards, Robert Schoonhoven From research <@t> nhls.ac.za Mon Oct 17 09:58:13 2005 From: research <@t> nhls.ac.za (Research) Date: Mon Oct 17 09:59:41 2005 Subject: [Histonet] C4d and SV 40 Message-ID: Hi all, Does anyone know who distributes these antibodies in South Africa: C4d - renal allograft biopsies - frozen tissue SV 40 - Bk polyoma virus infection Thanks in advance! S. Kahn Dept. Anatomical Pathology Johannesburg South Africa ********************************************************************************** The views expressed in this email are, unless otherwise stated, those of the author and not those of the National Health Laboratory Services or its management. The information in this e-mail is confidential and is intended solely for the addressee. Access to this e-mail by anyone else is unauthorized. If you are not the intended recipient, any disclosure, copying, distribution or any action taken or omitted in reliance on this, is prohibited and may be unlawful. Whilst all reasonable steps are taken to ensure the accuracy and integrity of information and data transmitted electronically and to preserve the confidentiality thereof, no liability or responsibility whatsoever is accepted if information or data is, for whatever reason, corrupted or does not reach its intended destination. *********************************************************************************** From nakagawa <@t> umn.edu Mon Oct 17 10:02:37 2005 From: nakagawa <@t> umn.edu (yasushi nakagawa) Date: Mon Oct 17 10:02:52 2005 Subject: [Histonet] xylene substitutes Message-ID: Hi all, I am wondering how good xylene replacements are compared with xylene. We use mouse brain sections (mostly 20um-thick cryosections, and some thicker sledge sections) and do immunostaining with cy-2/-3/-5 as well as Alexafluor conjugated secondary antibodies, GFP/RFP/CFP detection, and DAB reaction/Nissl staining. I wonder if someone knows long-term signal (color and fluorescence) retention and tissue quality when you use xylene substitutes before DPX mounting. I find some in EM Sciences catalog and would like to ask you if you have recommendations for particular products. Thank you. Yasushi Nakagawa Department of Neuroscience Stem Cell Institute University of Minnesota From rjbuesa <@t> yahoo.com Mon Oct 17 10:21:47 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Mon Oct 17 10:21:56 2005 Subject: [Histonet] xylene substitutes In-Reply-To: Message-ID: <20051017152147.75401.qmail@web61213.mail.yahoo.com> If you are looking for xylene substitutes because you are trying to avoid the chemical hazard that xylene has, all other xylene substitutes are also hazardous in some level and none work as well as xylene. If you are trying to avoid the hazard, you have to eliminate aromatic substances altogether and go for an aliphatic reagent. Since attachments are supposed not to be posted in Histonet I am sending a procedure directly to your e-mail address. This procedure retains fluorescence signals also. Rene J. yasushi nakagawa wrote: Hi all, I am wondering how good xylene replacements are compared with xylene. We use mouse brain sections (mostly 20um-thick cryosections, and some thicker sledge sections) and do immunostaining with cy-2/-3/-5 as well as Alexafluor conjugated secondary antibodies, GFP/RFP/CFP detection, and DAB reaction/Nissl staining. I wonder if someone knows long-term signal (color and fluorescence) retention and tissue quality when you use xylene substitutes before DPX mounting. I find some in EM Sciences catalog and would like to ask you if you have recommendations for particular products. Thank you. Yasushi Nakagawa Department of Neuroscience Stem Cell Institute University of Minnesota _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From pathrm35 <@t> adelphia.net Mon Oct 17 10:22:47 2005 From: pathrm35 <@t> adelphia.net (pathrm35@adelphia.net) Date: Mon Oct 17 10:22:57 2005 Subject: [Histonet] AB PAS and PASF CPT codes Message-ID: <23850347.1129562567613.JavaMail.root@web5.mail.adelphia.net> Fellow techs, What are the CPT codes for PAS for fungus (88312?)and Alcian Blue PAS? Also, what are the differences in reimbursements (money wise)? Thanks, Ron Martin From pathrm35 <@t> adelphia.net Mon Oct 17 10:33:29 2005 From: pathrm35 <@t> adelphia.net (pathrm35@adelphia.net) Date: Mon Oct 17 10:33:37 2005 Subject: [Histonet] CAP IHC disclaimer on path reports Message-ID: <12599435.1129563209733.JavaMail.root@web5.mail.adelphia.net> Fellow techs, What are you placing on your path reports reguarding disclaimers for IHC? This is reguarding CAP inspection ANP.12425, Class I analyte- specific reagents (antibodies). Please e mail me an example if possible. Thanks, Ron Martin From STapper <@t> slhduluth.com Mon Oct 17 11:10:59 2005 From: STapper <@t> slhduluth.com (Tapper, Sheila) Date: Mon Oct 17 11:11:09 2005 Subject: [Histonet] Block cutting protocols Message-ID: <9A3C36FB3D39404398867DFE2A4B5DE70109679E@slhw2smail01.slhdomain.com> NEVER - For all of the obvious reasons. The opportunity for error is way too great. Sheila Tapper HT(ASCP) St. Luke's Hospital Duluth, MN -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Histology SLU Sent: Monday, October 17, 2005 9:05 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Block cutting protocols Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This communication is intended for the use of the person or entity to whom it is addressed and may contain information that is privileged and confidential, the disclosure of which is governed by applicable law. If the reader of this message is not the intended recipient, or the employee or agent responsible for delivering it to the intended recipient, you are hereby notified that any dissemination, distribution, or copying of this information is prohibited. If you have received this message in error, please notify sender immediately. From sharon.osborn <@t> dnax.org Mon Oct 17 11:37:25 2005 From: sharon.osborn <@t> dnax.org (Osborn, Sharon) Date: Mon Oct 17 11:40:57 2005 Subject: [Histonet] licensing in CA Message-ID: <29B25753F6B1D51196110002A589D4440239823C@PALMSG30.us.schp.com> Linda, Currently, histology does not have a state licensing requirement. The only requirements for ASCP, degree, experience are imposed by the hiring institution. It is generally understood or required that someone taking a position will become certified within a certain period of time. sharon osborn DNAX, SP BioPharma Palo Alto, CA From: Linda Hines Subject: [Histonet] licensing for histology technician in california To: histonet@lists.utsouthwestern.edu Message-ID: <20051015211926.74502.qmail@web33005.mail.mud.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Hi! All I would like to know if there is a license requirement to work in california for histologists. I have been attempting to locate on line, no luck... do any of you that work in california know who to contact? If it is necessary? Thanks in advance. Everyone have a good week-end. Linda Hines HT 602-439-4104 Histo-Techs ON CALL ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. From relia1 <@t> earthlink.net Mon Oct 17 12:26:52 2005 From: relia1 <@t> earthlink.net (Pam Barker) Date: Mon Oct 17 12:27:03 2005 Subject: [Histonet] RELIA's Histology Job Opportunity Update 10/17/05 Message-ID: Hello Histonetters, I just wanted to drop you a line and tell you about my current histology openings. All the positions I represent are permanent positions with great companies who offer excellent compensation, benefits and relocation/sign on bonuses. Wherever you want to go, wherever you want to stay if you are looking for a new opportunity I can help. I offer over 20 years of recruiting experience and a recruiting practice solely dedicated to permanent placement in the Histology profession. In addition to representing you to the positions you are interested in with my clients I offer to you FREE of Charge: ? Assistance with updating or creating your resume ? Tips on interviewing ? Encouragement and assistance during the course of your job search ? Complete Confidentiality ( I will only represent you to jobs you tell me you are interested in looking into) Here is a list of my newest openings 1. Senior Research Specialist Immunohistochemistry - California 2. Histo Tech (2 positions) Clinical Setting - SE Georgia 3. Histo Tech (2 positions) Clinical Setting - SW Florida (eligibility for FL lic req) 4. Technical Support Specialist - Research Setting - Northern California 5. Capital Equipment Sales ? Multiple U.S. locations 6. Histo Tech ? Clinical Setting (Northern CA/Southern OR) Here is a list of my current openings: HISTOLOGY MANAGEMENT: 1. Anatomic Pathology Manager ? Boston, MA 2. Histology Supervisor ? Boston, MA 3. Histology Supervisor ? South Florida HISTOTECHNICIAN/TECHNOLOGIST: 1. Histo Tech (clinical setting) ? Northern, CA 2. Histo Tech (research setting) ? Northern, CA 3. Histo Tech (clinical setting) ? Central Florida 4. Histo Tech (clinical setting) - Southeastern Florida 5. Histo Tech (clinical setting) ? Boston, MA 6. Histo Tech (clinical setting) - Minnesota 7. Histo Tech (clinical setting) ? Alaska If you or any of your friends would like more information on any of these positions. Or if you would like to discuss opportunities in other areas or future job searches please contact me. I would be more than happy to assist you. You can reach me at 866-607-3542 or relia1@earthlink.net I work with facilities nationwide and I will keep your resume confidential. I will only represent you to jobs you tell me you are interested in looking into. Remember It never hurts to keep an eye open even if you are happy in your present job. Thank you, Pam ? 866-607-3542 (866-60RELIA) Thank You! Pam Barker President RELIA Specialists in Allied Healthcare Recruiting 5703 Red Bug Lake Road #330 Winter Springs, FL 32708-4969 Phone: (407)657-2027 Cell: (407)353-5070 FAX: (407)678-2788 E-mail: relia1@earthlink.net From lfidgen <@t> vt.edu Mon Oct 17 12:38:17 2005 From: lfidgen <@t> vt.edu (Laura Fidgen) Date: Mon Oct 17 12:38:28 2005 Subject: [Histonet] cryostats Message-ID: <6.0.0.22.0.20051017133658.026b1c38@pop.vt.edu> We are in the market for a new cryostat. Recommendations/suggestions would be greatly appreciated! Laura L. Fidgen, MScF, BSc, HT(ASCP) Medical Technologist VMRCVM, Histopatholgy Blacksburg, VA 24060-0443 From PIXLEYSK <@t> UCMAIL.UC.EDU Mon Oct 17 12:46:02 2005 From: PIXLEYSK <@t> UCMAIL.UC.EDU (Pixley, Sarah (pixleysk)) Date: Mon Oct 17 12:46:09 2005 Subject: [Histonet] Re: Antigen retrieval with anti-BrdU Message-ID: Dear Steve: I believe that your nuclear stain may come from the anti-BrdU and your detection system rather than your HIER method. Here are the results from one of my experiments. You can see that the nuclear staining went away under two conditions: diluting the primary or diluting the secondary and ABC. These were cryostat sectioned slices of mouse noses. The mice had been perfused with 4% paraformaldehyde, treated with formic acid for decalcification, then placed in a cryoprotectant containing sucrose, then frozen and sectioned. After air drying frozen sections, they were postfixed in 4% para for 10 mins, then 10 mins in hydrogen peroxide in 100% methanol, then treated with protease K, then blocked with 10% fetal calf serum/0.2% triton X-100 in PBS. The primary antibody was a sheep anti-BrdU (overnight at room temp in a humid chamber), the secondary was a biotinylated rabbit anti-sheep (2 hrs room temp) and the ABC was the Elite version (2 hrs rm temp). No counterstain was used. Slide # Slide Label 1?Ab 2?Ab ABC Results 2 1/1000 1/100 1/200 Good label but some Bkg in all nuclei. More Bkg than slide 5 3 1/5000 1/100 1/200 Beautiful! Dark Cells. No Bkg. Almost can't see tissue 4 1/1000 1/100 1/500 Good label. Has nuclear Bkg, but lighter than in slide 2 5 1/1000 1/400 1/200 Good label. More Bkg. Than in slide 4 (prob due to ABC) 6 1/1000 1.400 1/500 Very clean-No nuclear stain Bold are the concentrations I chose for further exps. Good luck. Sarah Pixley, Has anyone had problems with unwanted specific nuclear staining after performing heat AR especially from the microwave. I've been attempting to stain paraffin embedded rat liver in which a 24hour and 48hour BDL with Brdu injection 1 hour prior to harvest. With any form of AR, even limited 80C I get 75-100% nuclear staining. With no AR I get nothing, not even the bile ducts. Does AR tend to effect nuclear staining with some antibodies. I'm stumped. Steve From bjdewe <@t> aol.com Mon Oct 17 12:48:35 2005 From: bjdewe <@t> aol.com (bjdewe@aol.com) Date: Mon Oct 17 12:48:51 2005 Subject: [Histonet] cryostats In-Reply-To: <6.0.0.22.0.20051017133658.026b1c38@pop.vt.edu> References: <6.0.0.22.0.20051017133658.026b1c38@pop.vt.edu> Message-ID: <8C7A15A21CEADCA-15C8-148CD@MBLK-M01.sysops.aol.com> I'd go with one of the Microm HM 355 with the vacuum attachment...nice machine ;-) Lorie ******************************************************************* Confidentiality Notice: This e-mail message, including any attachments, is for the sole use of the intended recipient(s)and may contain confidential and privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by reply e-mail and destroy all copies of the original message. ******************************************************************* -----Original Message----- From: Laura Fidgen To: histonet@lists.utsouthwestern.edu Sent: Mon, 17 Oct 2005 13:38:17 -0400 Subject: [Histonet] cryostats We are in the market for a new cryostat. Recommendations/suggestions would be greatly appreciated! Laura L. Fidgen, MScF, BSc, HT(ASCP) Medical Technologist VMRCVM, Histopatholgy Blacksburg, VA 24060-0443 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From bjdewe <@t> aol.com Mon Oct 17 13:00:29 2005 From: bjdewe <@t> aol.com (bjdewe@aol.com) Date: Mon Oct 17 13:00:45 2005 Subject: [Histonet] cryostats In-Reply-To: <07AB60D5D7B9754EBF56F360F98D083DEB444D@fh2k093.fhmis.net> References: <07AB60D5D7B9754EBF56F360F98D083DEB444D@fh2k093.fhmis.net> Message-ID: <8C7A15BCB66CBCC-15C8-14AD2@MBLK-M01.sysops.aol.com> Of course, sorry...got my numbers switched ;-) HM 550... http://www.rallansci.com/instrumentation/instrumentation.aspx?id=144 Lorie ******************************************************************* Confidentiality Notice: This e-mail message, including any attachments, is for the sole use of the intended recipient(s)and may contain confidential and privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by reply e-mail and destroy all copies of the original message. ******************************************************************* -----Original Message----- From: Bonner, Janet To: 'bjdewe@aol.com ' ; 'histonet-bounces@lists.utsouthwestern.edu ' ; 'Histonet@lists.utsouthwestern.edu ' Sent: Mon, 17 Oct 2005 13:53:54 -0400 Subject: RE: [Histonet] cryostats Too bad the HM355 is a microtome, not a cryostat - @:) -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu To: Histonet@lists.utsouthwestern.edu Sent: 10/17/2005 1:48 PM Subject: Re: [Histonet] cryostats I'd go with one of the Microm HM 355 with the vacuum attachment...nice machine ;-) Lorie ******************************************************************* Confidentiality Notice: This e-mail message, including any attachments, is for the sole use of the intended recipient(s)and may contain confidential and privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by reply e-mail and destroy all copies of the original message. ******************************************************************* -----Original Message----- From: Laura Fidgen To: histonet@lists.utsouthwestern.edu Sent: Mon, 17 Oct 2005 13:38:17 -0400 Subject: [Histonet] cryostats We are in the market for a new cryostat. Recommendations/suggestions would be greatly appreciated! Laura L. Fidgen, MScF, BSc, HT(ASCP) Medical Technologist VMRCVM, Histopatholgy Blacksburg, VA 24060-0443 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From vazquezr <@t> ohsu.edu Mon Oct 17 13:04:33 2005 From: vazquezr <@t> ohsu.edu (Robyn Vazquez) Date: Mon Oct 17 13:04:57 2005 Subject: [Histonet] Block cutting protocols Message-ID: This is an excellent question. I would never put more than one patient on the water bath. Too many things can happen and you are setting yourself up to mix patients up. This is the only time a patient is NOTdentified, is on the water bath. Yes, you can compare blocks to match and that is a waste of time, what if two are so Identical, that you couldn't tell. To me it is UNETHICAL to do this. I worked with someone that did this and it made me just sick to see it happen. Supervisor was informed and she did nothing. Robyn OHSU From Myri37 <@t> aol.com Mon Oct 17 13:36:01 2005 From: Myri37 <@t> aol.com (Myri37@aol.com) Date: Mon Oct 17 13:36:17 2005 Subject: [Histonet] need training in histology Message-ID: <145.4fdcbe4e.30854911@aol.com> Hi histonetters, First, let me inform you that your website has always been a great help, consulting you has become indispensable. I am a technician in a tissue engineering lab. I end up doing histology in paraffin and resin for the past three years which I enjoyed very much. However?.: Histology in paraffin did not cause me any problem in the other hand the one in resin (methylmetacrylate.spurr and epon) cause me few complications considering the fact that my samples are implants of titanium coated with a layer of hydroxyapatite that?s also covered by a cellular layer rich in collagen fibers. I found problems in the choice of resin , the resin MMA and epon present an important retraction , I make thick sections of 100 um then grind them. But I can't make all of the possible staining for collagen and the tissue newly mineralized I would like to know if you know any information on specialize training in histology as close as possible to my field in another word the inclusion of hard resin of diferent biomaterial covered with tissue. the choice of the appropriate fixative, the choice of dehydratation solvent, the choice of resin the best timing for it, a good interpretation of the microscopic blades and make the difference between coloration artifact and reel element. Could you provide me with title of manuals that will help me answer some of these questions? Or where I can find them? And also if there are courses or training to follow? Thank you very much for your information Myriam B France From sjchtascp <@t> yahoo.com Mon Oct 17 14:52:08 2005 From: sjchtascp <@t> yahoo.com (Steven Coakley) Date: Mon Oct 17 14:52:16 2005 Subject: [Histonet] 4N HCL Message-ID: <20051017195208.98784.qmail@web90209.mail.scd.yahoo.com> I haven't had to figure normality for ages. would a 4N HCL solution be 4 times the HCL strength of a 1N HCL solution? Or is that to simple. Styeve --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From asmith <@t> mail.barry.edu Mon Oct 17 15:02:23 2005 From: asmith <@t> mail.barry.edu (Smith, Allen) Date: Mon Oct 17 15:03:21 2005 Subject: [Histonet] 4N HCL Message-ID: <5D2189E74151CC42BEC02906BA8996322B9115@exchsrv01.barrynet.barry.edu> Yes. 4N is 4 times as strong as 1N. Allen A. Smith, Ph.D. Professor of Anatomy Barry University School of Graduate Medical Sciences Podiatric Medicine and Surgery Miami Shores, Florida 33161 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Steven Coakley Sent: Monday, October 17, 2005 3:52 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] 4N HCL I haven't had to figure normality for ages. would a 4N HCL solution be 4 times the HCL strength of a 1N HCL solution? Or is that to simple. Styeve --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet The information transmitted is intended only for the person or entity to which it is addressed and may contain confidential, and/or privileged material. No confidentiality or privilege is waived or lost by any errant transmission. If you receive this message in error, please immediately delete it and all copies of it from your system and notify the sender. E-mail transmission cannot be guaranteed to be secure or error-free as information could be intercepted, corrupted, lost, destroyed, arrive late or incomplete, or contain viruses. Barry University - Miami Shores, FL (http://www.barry.edu) From MICHAEL.OWEN <@t> fda.gov Mon Oct 17 15:15:35 2005 From: MICHAEL.OWEN <@t> fda.gov (Owen, Michael P) Date: Mon Oct 17 15:15:59 2005 Subject: [Histonet] 4N HCL Message-ID: Dear Steven, The information below combined with the recent post should give you some assistance. Good luck. Center for Food Safety and Applied Nutrition Bacteriological Analytical Manual Online January 2001 Reagents Index R36 1 N Hydrochloric Acid HCl (concentrated) 89 ml Distilled water to make 1 liter Based on this data, 4 N HCl would be prepared by adding (89 * 4 = 356 mL) of concentrated HCl to 644 mL distilled water. Remember to add acid to water for safety. I highly recommend you place the container that will hold the solution into an tub of ice or ice water when you combine the acid and water as the reaction is very exothermic. Michael P. Owen, Regulatory Microbiologist U.S. Food and Drug Administration Pacific Regional Lab Northwest 22201 23rd Drive SE Bothell, WA 98021-4421 Phone: 425-483-4865 E-Mail: michael.owen@fda.gov From PMonfils <@t> Lifespan.org Mon Oct 17 15:22:33 2005 From: PMonfils <@t> Lifespan.org (Monfils, Paul) Date: Mon Oct 17 15:22:51 2005 Subject: [Histonet] 4N HCL Message-ID: <09C945920A6B654199F7A58A1D7D1FDE017175F5@lsexch.lsmaster.lifespan.org> Since the normality of commercial HCl is (approximately) 12, you would dilute the commercial product 1:11 for a (approximately) 1N solution (final volume = 12x the original volume), or 1:2 (final volume = 3x the original volume) for a (approximately) 4N solution. > ---------- > From: histonet-bounces@lists.utsouthwestern.edu on behalf of > Steven Coakley > Sent: Monday, October 17, 2005 12:52 PM > To: Histonet@lists.utsouthwestern.edu > Subject: [Histonet] 4N HCL > > I haven't had to figure normality for ages. would a 4N HCL solution be 4 > times the HCL strength of a 1N HCL solution? Or is that to simple. > > Styeve > > > --------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > From tpmorken <@t> labvision.com Mon Oct 17 15:30:56 2005 From: tpmorken <@t> labvision.com (Morken, Tim - Labvision) Date: Mon Oct 17 15:31:13 2005 Subject: [Histonet] 4N HCL Message-ID: <0556BE8AC5551E4E8AF6BB9E42509BA20328D248@usca0082k08.labvision.apogent.com> Although histology and old chemistry books are full of Normal solutions, it is an out-of-date term as far as I can tell. HCl is easy because it has only one valence, but if you have more than one valency, with two or more ions with different valences, it becomes complicated as to what "normal" means ("normal" to what?). A more clear method is to express it in terms of equivilent molarity. An note about the use of "Nomal" soutions: http://www.madsci.org/posts/archives/2005-02/1108587746.Ch.r.html Comment is reproduced below: "Anyone who learnt any chemistry after about 1970 has probably never heard of a "gram equivalent weight" nor of the "normality" of a solution. But this is what Webster's definition a. is referring to. These terms were extensively used 50 years ago and before. In modern terms, the gram equivalent weight of a substance is equal to the molar mass divided by the valency. A 1 normal solution of hydrochloric acid is exactly the same as a 1 molar solution of hydrochloric acid, but a 1 normal solution of sulfuric acid is the same as a 0.5 molar solution of sulfuric acid (because the 'valency' of sulfuric acid is 2). The problem with normality as a measure of concentration was that the normality of the same solution could vary depending on the context of what you wanted to do with it. For example, dilute nitric acid has a 'valency' of 1 as an acid, but a 'valency' of 3 as an oxidant. So a solution of nitric acid that was 1 normal for the purpose of neutralizing with caustic soda was 3 normal for the purpose of dissolving up copper foil. Sometime in the 1960s, the International Union of Pure and Appplied Chemistry wisely decided that normality was a concept that chemists would be better off without, and that thenceforth concentrations would be expressed in terms of molarity. The bottom line as far as your question is concerned is that there are two possibilities. If the reference is to sodium chloride as 'normal saline' in a medical context, it means 155 millimolar; if, on the other hand it is a very old-fashioned chemical reference, then it means 1 molar, because the 'valency' of sodium chloride in any context is 1. " And this from International Union of Pure and Applied Chemistry (www.iupac.org) Seet their nomenclature book http://www.iupac.org/publications/analytical_compendium/ Chapter six covers equivilent solutions: ( www.iupac.org/publications/analytical_compendium/Cha06sec3.pdf ) Note the last sentence on page 6-5. (paragraph reproduced below) Normal solution A solution in which the amount-of-substance concentration of the equivalent of the reagent is1 mol dm-3(i.e. 1 mol l-1) was termed a Normal solution, symbol N. Decimalised fractions of N was used, e.g. 0.326 N H2SO4, that is a solution with c(1/2H2SO4) = 0.326 mol l-1. Note: Because confusion may exist when a reagent has different equivalence factors according to circumstances, the statements of normality must be accompanied by the equivalence factor, e.g.0.1 N KIO3; feq(KIO3) = 1/60.05 N KIO3; feq(KIO3) = 1/4. The use of the terms "Normal solution, Normality" are not recommended. Tim Morken Lab Vision - Neomarkers www.labvision.com Free webhosting for US State Histotechnology Societies: http://www.labvisioncorp.com/demowebsite/index.cfm -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Steven Coakley Sent: Monday, October 17, 2005 3:52 PM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] 4N HCL I haven't had to figure normality for ages. would a 4N HCL solution be 4 times the HCL strength of a 1N HCL solution? Or is that to simple. Styeve From PMonfils <@t> Lifespan.org Mon Oct 17 15:44:16 2005 From: PMonfils <@t> Lifespan.org (Monfils, Paul) Date: Mon Oct 17 15:44:28 2005 Subject: [Histonet] Block cutting protocols Message-ID: <09C945920A6B654199F7A58A1D7D1FDE017175F6@lsexch.lsmaster.lifespan.org> While it doesn't really address the original question, another related rule we strictly enforce in our laboratory, for exactly the same reasons, is - only one cassette may be open on the embedding unit at any one time. A tech may take four or five cassettes and place them on the working surface of the unit together, but only one cassette may be opened, and that cassette and its contents must be removed from the working surface of the unit before another cassette is opened. I assume most labs have a similar rule in place. > ---------- > From: histonet-bounces@lists.utsouthwestern.edu on behalf of > Histology SLU > Sent: Monday, October 17, 2005 7:04 AM > To: histonet@lists.utsouthwestern.edu > Subject: [Histonet] Block cutting protocols > > Hello All: > > I hope that you all won't "flame" me with this question. I do not want to > offer any information yet as to why I am asking. I want your responses to > be unbiased by the circumstances for which I am asking. > > The question: > > Do any of you techs cutting clinical paraffin embedded blocks, lay out > multiple ribbons from multiple blocks on your water bath at one time and > then pick up your sections for each block? > > I would like tech and supervisor responses please. Thanks so much. You > guys are always such a help. > > Susan > > > --------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > From ohenry <@t> dfw.net Mon Oct 17 16:16:52 2005 From: ohenry <@t> dfw.net (Susan Owens) Date: Mon Oct 17 16:17:14 2005 Subject: [Histonet] Re:Block cutting protocols Message-ID: <000701c5d360$1ce1d310$62dd3040@your4f1261a8e5> To lay out multiple ribbons from different blocks on the water bath at the same time IS A DISASTER LOOKING TO HAPPEN. It should never be allowed. Susan Owens-Tx >Date: Mon, 17 Oct 2005 07:04:43 -0700 (PDT) >Subject: [Histonet] Block cutting protocols >Hello All: >I hope that you all won't "flame" me with this question. I do not want to >offer any information yet as >to why I am asking. I want your responses to >be unbiased by the circumstances for which I am >asking. >The question: >Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple >blocks on your water bath at one time and then pick up your sections for each block? >I would like tech and supervisor responses please. Thanks so much. You >guys are always such a >help. >Susan From ralphpu <@t> alleninstitute.org Mon Oct 17 20:05:00 2005 From: ralphpu <@t> alleninstitute.org (Ralph Puchalski) Date: Mon Oct 17 20:05:12 2005 Subject: [Histonet] Pink precipitate is monoformazan from BCIP/NBT reaction onISH? Message-ID: Dear John, Thank you for your advice. Yes, we use digoxigenin (DIG)-labeled RNA probes (riboprobes) in the in situ hybridization of frozen sections to map gene expression throughout the mouse brain. Detection of bound riboprobe is a multi-step procedure. First, a succession of blocking steps inhibits endogenous protein activity from interfering with the colorimetric enzymatic reactions. The colorimetric reaction itself is a four part process, starting with the addition of a peroxidase-conjugated antibody directed at the DIG-UTP hapten incorporated in the bound riboprobe. A Tyramide Signal Amplification step is utilized to maximize sensitivity. In brief, biotin-coupled tyramide is added to the tissue, resulting in the formation of multiple activated tyramide molecules through the activity of each bound peroxidase-coupled antibody molecule. These highly reactive tyramide radicals bind to protein residues in the vicinity of the bound riboprobe, thereby resulting in an amplification of bound biotin molecules available for detection by up to a hundred fold (relative to the number of bound antibody molecules). These biotin molecules are then bound to NeutrAvidin-AP, the third step of the colorimetric reaction. Alkaline phosphatase (AP) conjugated to the NeutrAvidin (NA-AP) enzymatically cleaves the phosphate from 5-bromo-4-chloro-3-indolyl phosphate (BCIP), and two of the resulting indoles undergo a redox reaction with nitroblue tetrazolium (NBT) to produce a blue particulate precipitate at the sites of riboprobe binding. We process about 800 slides (1600 sections per day), and you can visit our site for more details of our method and product at: http://www.brain-map.org/pdf/ABADataProductionProcesses.pdf;jsessionid=9 FF15AEC860CC06CC8ECD7E42FF46402 Processing 800 slides per day in order to cover the entire genome of about 20,000 genes necessitates the use of robots. The slides are incubated in only 300 ul of a particular reagent solution in chambers mounted on an angle. The incubation periods and concentrations were optimized for detection of the lowest levels of target RNA expression. The conditions were established for the entire project and major changes cannot be made. The system was set up so that all sections receive the same amount of all reagents, including BCIP, NBT, and NA-AP. You commented on nothing dehydrogenases. Our experiments demonstrate that removing NA-AP from the reaction, while keeping all other steps as they are in the control, produces sections that are transparent, i.e. without blue or purple staining under bright-field microscopy. If anything, the sections have a very faint yellow tint, the same as what we observe in the bottles just after we prepare the BCIP/NBT substrate in buffer with Levamisole. However, if we incubate these slides in a reducing agent such as 50 mM alkaline ascorbate for 5 minutes in pH 9.5 in Tris-Magnesium-Saline, the slides paired to the ones that are transparent become stained deep purple-blue (no NA-AP). Therefore, if we are interpreting the data correctly, nothing dehydrogenases contribute nothing that is detectable, and ascorbate reduces NBT to diformazan. Do you agree? The troubling point is that the amount of staining visualized by reduction with ascorbate matches the staining by the pink precipitate (formed in the presence of NA-AP in our production runs and controls), which presumably is monoformazan as determined spectrophotometrically. The staining levels are highest in the fiber tracts, brainstem, and hypothalamus, but are visible throughout the sections to various extents. The reason why this is troubling is that the staining does not require NA-AP. The NBT (we will test to be sure BCIP is not required) just sticks to these areas, and later ends up on production slides (NA-AP+) as monoformazan or pink precipitate, which is our hypothesis. Do you agree with this assessment? How would you go about decreasing the staining of the fiber tracts, brainstem, and hypothalamus, and the rest of the section with NBT? We plan to titrate the NBT, but cannot reduce signal levels. Our fixation and dehydration process does not remove all the fats in the tissue, so it is quite possible that the hydrophobic NBT gets partitioned into the lipid phase. Would you attempt to change the BCIP/NBT/Levamisole buffer (Tris-HCl, NaCl, and MgCl2, Tween 20) used in the incubation with NA-AP for color development? Do you know if anyone has tried to block the endogenous sites to which NBT adheres? Without knowing how the binding occurs, it is difficult to suggest options. How would you go about characterizing the binding of NBT to the sections? I wish it were as easy as finding a convenient analogue of NBT that is colorless, does not inhibit AP activity, is not reduced, and binds to the same sites as NBT does, thereby blocking NBT from binding to these sites. Any suggestions? Your advice is appreciated. Ralph Puchalski -----Original Message----- From: John Kiernan [mailto:jkiernan@uwo.ca] Sent: Monday, September 26, 2005 9:42 PM To: Ralph Puchalski Cc: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Pink precipitate is monoformazan from BCIP/NBT reaction onISH? Dear Ralph, Your email is full of abbreviations and jargon. Am I right in thinking it's a question about localizing alkaline phosphatase activity by an indoxyl-tetrazolium method? If so, please provide a reference to the original publication and let us all know if you followed it exactly or made any changes. Part of your email suggests that the alkaline phosphatase activity is not endogenous but part of an amplification system used in in situ hybridization. The pH optima of endogenous and label alkaline phosphatases differ. The later paragraphs of your message indicate that you may not fully understand the significance of the mono- and diformazan products of reduction of nitro-BT. Both colours result from reduction of the tetrazolium salt, but you need controls to prove that the reduction was by bromochloroindoxyl and not by other reducing agents (such as diaphorases) in the tissue. Non-enzymatic reduction of tetrazolium salts by -SH has been a well known artifact {"nothing dehydrogenase") for 40-45 years. John A. Kiernan Anatomy Dept, UWO London, Canada. ___________________________________________________ Ralph Puchalski wrote: > > I am trying to figure out the origin of the pink precipitate in the > image I posted on http://www.histonet.org/site_images_frame.asp > > Please go to the image entitled: Pink Precipitate ver2.jpg. It is at > the top of the list on 9/26/05, posted by Ralph Puchalski. To see the > pink precipitate artifact, please open the image and set the scroll bars > on the bottom and right side of the image at their 1/2 way points. It > is ugly! > > I think this precipitate is the aggregation of the monoformazan > intermediate generated from NBT (after dephosphorylation of BCIP by > alkaline phosphatase) that is not completely reduced to diformazan, the > insoluble dark blue or black precipitate that labels cells expressing > target mRNAs in our in situ hybridization reactions. > > We don't know how the monoformazan adheres to the sections of tissue. > It appears to be non-covalent due to the tendency of the monoformazan to > migrate under the coverslip in the aqueous mounting medium that has yet > to dry, and form clumps or aggregates or pools as seen in the picture. > The monoformazan is soluble in ethanol so doesn't form pools or > aggregates. But as soon as it is exposed to an aqueous medium, it > precipitates. > > If we mount the post-ISH tissue sections with an organic based mounting > medium like UV-CureMount (Instrumedics), the monoformazan might cause > the entire section to turn to a brown tint as seen on > http://www.histonet.org/site_images_frame.asp > > Please go to the image Brown Tint 1.jpg. It is 4th image from the top > on 9/26, posted by Ralph Puchalski. The lower image is mounted with UV > CureMount, and the upper was with aqueous Hydro Matrix. There is no > pooling or precipitation of monoformazan with UV CureMount, but I think > it does cause the entire section to turn brown. > > Question: How do we eliminate this problem, which I believe is > monoformazan? If we reduce it fully using ascorbate, the section (later > mounted with HyrdoMatrix) turns dark blue or purple, the same color as > our ISH signal. We have tried washing off the monoformazan with 100% > ethanol prior to coverslipping, but only small amounts are removed. > 100% acetone also does not work effectively. > > Please let me know if you have any ideas that might help us eliminate > this problem. > > Thank you, > > Ralph > > Ralph Puchalski, Ph.D. > Manager, Process Engineering and Automation > Allen Institute for Brain Science > 551 N. 34th Street, Suite 200 > Seattle, WA 98103 > > ralphpu@alleninstitute.org > Tel: 206-548-7041 Fax: 206-548-7071 > www.brainatlas.org > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From bills <@t> icpmr.wsahs.nsw.gov.au Mon Oct 17 20:15:40 2005 From: bills <@t> icpmr.wsahs.nsw.gov.au (Bill Sinai) Date: Mon Oct 17 20:16:32 2005 Subject: [Histonet] Alcian yellow Message-ID: <108A4A90116@icpmr.wsahs.nsw.gov.au> Does anyone know of a company who still manufactures Alcian Yellow??? Thanks Bill Sinai Laboratory Manager Tissue Pathology ICPMR Westmead Hospital Westmead NSW 2145 Phone 02 9845 7774 Fax 02 9687 2330 __________________________________________________________________ This electronic message and any attachments may be confidential. If you are not the intended recipient of this message would you please delete the message and any attachments and advise the sender. Western Sydney Area Health Services (WSAHS) uses virus scanning software but excludes any liability for viruses contained in any email or attachment. This email may contain privileged and confidential information intended only for the use of the addressees named above. If you are not the intended recipient of this email, you are hereby notified that any use, dissemination, distribution, or reproduction of this email is prohibited. If you have received this email in error, please notify WSAHS immediately. Any views expressed in this email are those of the individual sender except where the sender expressly and with authority states them to be the views of WSAHS. From BMolinari <@t> heart.thi.tmc.edu Tue Oct 18 05:43:55 2005 From: BMolinari <@t> heart.thi.tmc.edu (Molinari, Betsy) Date: Tue Oct 18 05:45:42 2005 Subject: [Histonet] Block cutting protocols Message-ID: No Betsy Molinari HT (ASCP) Texas Heart Institute Cardiovascular Pathology 6770 Bertner Ave. Houston,TX 77030 832-355-6524 832-355-6812 (fax) -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Histology SLU Sent: Monday, October 17, 2005 9:05 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Block cutting protocols Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From JPCOLEMA <@t> sentara.com Tue Oct 18 06:12:23 2005 From: JPCOLEMA <@t> sentara.com (JOHN COLEMAN) Date: Tue Oct 18 06:12:53 2005 Subject: [Histonet] ribbons on the water bath. Message-ID: About multiple patient ribbons on a bath at once- One word: NEVER From vanann702 <@t> skmc.gov.ae Tue Oct 18 06:41:00 2005 From: vanann702 <@t> skmc.gov.ae (Anne Van Binsbergen) Date: Tue Oct 18 06:39:48 2005 Subject: [Histonet] mercury Message-ID: <0C44F1AAEE47D54DA4210A60AB206F5E02F6D670@SKMCEMAIL.skmc.gov.ae> Do none of you remember the many lengthy discussions way back in the early days regarding Mercury and its use in Histo/AP labs???? There were even a few postings from Pathologists....come on guys ...I need your help here. Where are all the 'old' faces of the histonet? Who still uses it and why Who has stopped using it - why ....and how What is the most acceptable replacement - Zinc or good old NBF Any other comments welcome Have had a few direct replies. Thanks for those Others also would like to see so please also reply to histonet Hope to have that expected 'flurry' really soon. Cheers Annie From vanann702 <@t> skmc.gov.ae Tue Oct 18 06:41:00 2005 From: vanann702 <@t> skmc.gov.ae (Anne Van Binsbergen) Date: Tue Oct 18 06:40:06 2005 Subject: [Histonet] mercury Message-ID: <0C44F1AAEE47D54DA4210A60AB206F5E02F6D670@SKMCEMAIL.skmc.gov.ae> Do none of you remember the many lengthy discussions way back in the early days regarding Mercury and its use in Histo/AP labs???? There were even a few postings from Pathologists....come on guys ...I need your help here. Where are all the 'old' faces of the histonet? Who still uses it and why Who has stopped using it - why ....and how What is the most acceptable replacement - Zinc or good old NBF Any other comments welcome Have had a few direct replies. Thanks for those Others also would like to see so please also reply to histonet Hope to have that expected 'flurry' really soon. Cheers Annie From Emily.Wiesner <@t> medecine.unige.ch Tue Oct 18 09:15:41 2005 From: Emily.Wiesner <@t> medecine.unige.ch (Emily Jane Wiesner-Camm) Date: Tue Oct 18 09:16:20 2005 Subject: [Histonet] Storing frozen rat brain slices Message-ID: <4355038D.80606@medecine.unige.ch> Question: I was wondering if anyone could let me know what is the optimal storage for frozen rat brain slices (approx 2mm thick) which are in OCT: -20 degrees celsius or -80? Thanks in advance, Emily From froyer <@t> bitstream.net Tue Oct 18 09:29:51 2005 From: froyer <@t> bitstream.net (Ford Royer) Date: Tue Oct 18 09:30:06 2005 Subject: [Histonet] Leica 2155 Microtome Available In-Reply-To: <09C945920A6B654199F7A58A1D7D1FDE017175F6@lsexch.lsmaster.lifespan.org> Message-ID: <001e01c5d3f0$67109940$0b00a8c0@fords> I have been asked by a friend to post this on the HistoNet. His animal research lab is closing and he has an "almost new", used less than 9 months, Leica 2155 motorized microtome that he would like to sell... If anyone is interested, please contact me "Off the List" for more details and specifications. This could be a great opportunity for a real bargain. Thanks, ~ Ford Ford M. Royer, MT(ASCP) Minnesota Medical Specialists, Inc. 7177 Madison Ave. W. Golden Valley, MN 55427-3601 763-542-8725 phone 888-790-9686 toll free 763-546-4830 fax email From PMonfils <@t> Lifespan.org Tue Oct 18 10:12:21 2005 From: PMonfils <@t> Lifespan.org (Monfils, Paul) Date: Tue Oct 18 10:13:31 2005 Subject: [Histonet] Alcian yellow Message-ID: <09C945920A6B654199F7A58A1D7D1FDE017175F7@lsexch.lsmaster.lifespan.org> Do a search for the chemical at http://www.chemexper.com/ and you will probably find some suppliers. From cwscouten <@t> myneurolab.com Tue Oct 18 10:25:40 2005 From: cwscouten <@t> myneurolab.com (Charles Scouten) Date: Tue Oct 18 10:26:25 2005 Subject: [Histonet] Storing frozen rat brain slices Message-ID: <5784D843593D874C93E9BADCB87342AB91678F@tpiserver03.Coretech-holdings.com> Even -80 is not cold enough, but the colder the better. The ice will gradually restructure as crystalline ice, and expand. This will break cell membranes and disrupt the tissue quality. Use as early as possible. See the link: http://www.myneurolab.com/global/Manuals/Tips%20and%20Techniques%20Freez ing%20Artifact.pdf Cordially, Charles W. Scouten, Ph.D. myNeuroLab.com 5918 Evergreen Blvd. St. Louis, MO 63134 Ph: 314 522 0300 x 342 FAX 314 522 0377 cwscouten@myneurolab.com http://www.myneurolab.com -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Emily Jane Wiesner-Camm Sent: Tuesday, October 18, 2005 9:16 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Storing frozen rat brain slices Question: I was wondering if anyone could let me know what is the optimal storage for frozen rat brain slices (approx 2mm thick) which are in OCT: -20 degrees celsius or -80? Thanks in advance, Emily _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jkiernan <@t> uwo.ca Tue Oct 18 10:27:44 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Tue Oct 18 10:27:53 2005 Subject: [Histonet] Pink precipitate is monoformazan from BCIP/NBT reactiononISH? References: Message-ID: <43551470.3A05878E@uwo.ca> Dear Ralph, Eleven thoughts follow. 1. You state that "The incubation periods and concentrations were optimized for detection of the lowest levels of target RNA expression." Does this mean you initially had success with in situ hybridization, biotinylated tyramide amplification of the peroxidase label and further amplification by an avidin-alkaline phosphatase step and BCIP-nitroBT? 2. Were the riboprobe concentrations, times, temperatures etc chosen on the basis of some other method? If they were based on stained blots, especially if the hybridization signals were detected by a different method, the same conditions probably would not be optimal for sections of tissue. In a blot, the mRNA concentrations are very low because each band has an area of several square millimetres. In a section the local concentration in a cell that's busy expressing a gene is going to be much higher, with the expressing cell surrounded by other stuff (mostly parts of other cells , in brain tissue) that do not contain that mRNA. [Sorry for that very long sentence!] Your sections might be receiving too much riboprobe (nonspecific attachment and staining everywhere , albeit in the wrong colour. Alternatively the sections might be failing to bind any riboprobe, and the pink monoformazan is just a consequence of excessive incubation in the last stage of a ridiculously complicated method that has detected nothing. 3. NitroBT is a large lipophilic cation (Horobin 2002; Chapter 5, p.166 in the 10th edition of Conn's Biological Stains). As such, it can be expected to adhere more to white matter than to grey matter in either frozen or paraffin sections 4. When you deliberately reduced the nitroBT with ascorbate, in known hybridization-negative sections, you proved that (a) your alkaline phosphatase detection method could generate blue diformazan, (b) the natural disposition of the nitroBT was what you would expect from its well documented chemistry and histochemistry, and (c) you may not washing sections well enough to remove weakly bound reagents. 5. Have you seen localization of any well studied mRNA in a well documented site in the brain? If not, your procedure will never localize the sites of transcription or translation of any genes. 6. You state "First, a succession of blocking steps inhibits endogenous protein activity from interfering with ..." That's standard practice in immunohistochemistry and in situ hybridization, but the blocking reactions need ro be understood by the researcher. Some blocking methods can, if misused, provide wrong false-negative controls. 7. I haven't yet visited http://www.brain-map.org/pdf/ABADataProductionProcesses.pdf;jsessionid=9 FF15AEC860CC06CC8ECD7E42FF46402 to review the technical details of the methods carried out by your robots. I may have a look later in the week, but don't expect a detailed review by way of Histonet. 8. My guess is that your in situ hybridization technique, despite its multiple amplifications, is detecting nothing when applied to sections. 9. A possible explanation for the pink nothingness may be your "succession of blocking steps." Is there one that extracts mRNA or interferes with subsequent hybridization? Protein blocking procedures might prevent the oxidation products of biotinylated tyramine from binding to the tyrosine side-chains assumed to occur universally in animal tissues, 10. Before setting the robots to work on 20,000 genes, try to get the method to work under human control for 5 genes. If difficulties arise with even one of the five, there can be no justification for investing 4000 times as much money in applying the technology on a larger scale. 11. In every kind of histological or histochemical staining the chance of a fatal error increases with the number of steps in the technique and with the number of slides being processed. You need to get your method right for individual slides and then groups of 10, long before attempting to process 800 slides in a day. John Kiernan Anatomy, UWO London, Canada --------------------------- Ralph Puchalski wrote: > > Dear John, > > Thank you for your advice. Yes, we use digoxigenin (DIG)-labeled RNA > probes (riboprobes) in the in situ hybridization of frozen sections to > map gene expression throughout the mouse brain. Detection of bound > riboprobe is a multi-step procedure. First, a succession of blocking > steps inhibits endogenous protein activity from interfering with the > colorimetric enzymatic reactions. The colorimetric reaction itself is a > four part process, starting with the addition of a peroxidase-conjugated > antibody directed at the DIG-UTP hapten incorporated in the bound > riboprobe. A Tyramide Signal Amplification step is utilized to maximize > sensitivity. In brief, biotin-coupled tyramide is added to the tissue, > resulting in the formation of multiple activated tyramide molecules > through the activity of each bound peroxidase-coupled antibody molecule. > These highly reactive tyramide radicals bind to protein residues in the > vicinity of the bound riboprobe, thereby resulting in an amplification > of bound biotin molecules available for detection by up to a hundred > fold (relative to the number of bound antibody molecules). These biotin > molecules are then bound to NeutrAvidin-AP, the third step of the > colorimetric reaction. Alkaline phosphatase (AP) conjugated to the > NeutrAvidin (NA-AP) enzymatically cleaves the phosphate from > 5-bromo-4-chloro-3-indolyl phosphate (BCIP), and two of the resulting > indoles undergo a redox reaction with nitroblue tetrazolium (NBT) to > produce a blue particulate precipitate at the sites of riboprobe > binding. We process about 800 slides (1600 sections per day), and you > can visit our site for more details of our method and product at: > http://www.brain-map.org/pdf/ABADataProductionProcesses.pdf;jsessionid=9 > FF15AEC860CC06CC8ECD7E42FF46402 > > Processing 800 slides per day in order to cover the entire genome of > about 20,000 genes necessitates the use of robots. The slides are > incubated in only 300 ul of a particular reagent solution in chambers > mounted on an angle. The incubation periods and concentrations were > optimized for detection of the lowest levels of target RNA expression. > The conditions were established for the entire project and major changes > cannot be made. The system was set up so that all sections receive the > same amount of all reagents, including BCIP, NBT, and NA-AP. > > You commented on nothing dehydrogenases. Our experiments demonstrate > that removing NA-AP from the reaction, while keeping all other steps as > they are in the control, produces sections that are transparent, i.e. > without blue or purple staining under bright-field microscopy. If > anything, the sections have a very faint yellow tint, the same as what > we observe in the bottles just after we prepare the BCIP/NBT substrate > in buffer with Levamisole. However, if we incubate these slides in a > reducing agent such as 50 mM alkaline ascorbate for 5 minutes in pH 9.5 > in Tris-Magnesium-Saline, the slides paired to the ones that are > transparent become stained deep purple-blue (no NA-AP). Therefore, if > we are interpreting the data correctly, nothing dehydrogenases > contribute nothing that is detectable, and ascorbate reduces NBT to > diformazan. Do you agree? > > The troubling point is that the amount of staining visualized by > reduction with ascorbate matches the staining by the pink precipitate > (formed in the presence of NA-AP in our production runs and controls), > which presumably is monoformazan as determined spectrophotometrically. > The staining levels are highest in the fiber tracts, brainstem, and > hypothalamus, but are visible throughout the sections to various > extents. The reason why this is troubling is that the staining does not > require NA-AP. The NBT (we will test to be sure BCIP is not required) > just sticks to these areas, and later ends up on production slides > (NA-AP+) as monoformazan or pink precipitate, which is our hypothesis. > Do you agree with this assessment? > > How would you go about decreasing the staining of the fiber tracts, > brainstem, and hypothalamus, and the rest of the section with NBT? We > plan to titrate the NBT, but cannot reduce signal levels. Our fixation > and dehydration process does not remove all the fats in the tissue, so > it is quite possible that the hydrophobic NBT gets partitioned into the > lipid phase. Would you attempt to change the BCIP/NBT/Levamisole buffer > (Tris-HCl, NaCl, and MgCl2, Tween 20) used in the incubation with NA-AP > for color development? > > Do you know if anyone has tried to block the endogenous sites to which > NBT adheres? Without knowing how the binding occurs, it is difficult to > suggest options. How would you go about characterizing the binding of > NBT to the sections? I wish it were as easy as finding a convenient > analogue of NBT that is colorless, does not inhibit AP activity, is not > reduced, and binds to the same sites as NBT does, thereby blocking NBT > from binding to these sites. Any suggestions? > > Your advice is appreciated. > > Ralph Puchalski > > -----Original Message----- > From: John Kiernan [mailto:jkiernan@uwo.ca] > Sent: Monday, September 26, 2005 9:42 PM > To: Ralph Puchalski > Cc: histonet@lists.utsouthwestern.edu > Subject: Re: [Histonet] Pink precipitate is monoformazan from BCIP/NBT > reaction onISH? > > Dear Ralph, > > Your email is full of abbreviations and jargon. > Am I right in thinking it's a question about > localizing alkaline phosphatase activity by an > indoxyl-tetrazolium method? If so, please provide > a reference to the original publication and let us > all know if you followed it exactly or made any > changes. Part of your email suggests that the > alkaline phosphatase activity is not endogenous > but part of an amplification system used in > in situ hybridization. The pH optima of endogenous > and label alkaline phosphatases differ. > > The later paragraphs of your message indicate that > you may not fully understand the significance of > the mono- and diformazan products of reduction of > nitro-BT. Both colours result from reduction of > the tetrazolium salt, but you need controls to > prove that the reduction was by bromochloroindoxyl > and not by other reducing agents (such as diaphorases) > in the tissue. Non-enzymatic reduction of tetrazolium > salts by -SH has been a well known artifact {"nothing > dehydrogenase") for 40-45 years. > > John A. Kiernan > Anatomy Dept, UWO > London, Canada. > ___________________________________________________ > Ralph Puchalski wrote: > > > > I am trying to figure out the origin of the pink precipitate in the > > image I posted on http://www.histonet.org/site_images_frame.asp > > > > Please go to the image entitled: Pink Precipitate ver2.jpg. It is at > > the top of the list on 9/26/05, posted by Ralph Puchalski. To see the > > pink precipitate artifact, please open the image and set the scroll > bars > > on the bottom and right side of the image at their 1/2 way points. It > > is ugly! > > > > I think this precipitate is the aggregation of the monoformazan > > intermediate generated from NBT (after dephosphorylation of BCIP by > > alkaline phosphatase) that is not completely reduced to diformazan, > the > > insoluble dark blue or black precipitate that labels cells expressing > > target mRNAs in our in situ hybridization reactions. > > > > We don't know how the monoformazan adheres to the sections of tissue. > > It appears to be non-covalent due to the tendency of the monoformazan > to > > migrate under the coverslip in the aqueous mounting medium that has > yet > > to dry, and form clumps or aggregates or pools as seen in the picture. > > The monoformazan is soluble in ethanol so doesn't form pools or > > aggregates. But as soon as it is exposed to an aqueous medium, it > > precipitates. > > > > If we mount the post-ISH tissue sections with an organic based > mounting > > medium like UV-CureMount (Instrumedics), the monoformazan might cause > > the entire section to turn to a brown tint as seen on > > http://www.histonet.org/site_images_frame.asp > > > > Please go to the image Brown Tint 1.jpg. It is 4th image from the top > > on 9/26, posted by Ralph Puchalski. The lower image is mounted with > UV > > CureMount, and the upper was with aqueous Hydro Matrix. There is no > > pooling or precipitation of monoformazan with UV CureMount, but I > think > > it does cause the entire section to turn brown. > > > > Question: How do we eliminate this problem, which I believe is > > monoformazan? If we reduce it fully using ascorbate, the section > (later > > mounted with HyrdoMatrix) turns dark blue or purple, the same color as > > our ISH signal. We have tried washing off the monoformazan with 100% > > ethanol prior to coverslipping, but only small amounts are removed. > > 100% acetone also does not work effectively. > > > > Please let me know if you have any ideas that might help us eliminate > > this problem. > > > > Thank you, > > > > Ralph > > > > Ralph Puchalski, Ph.D. > > Manager, Process Engineering and Automation > > Allen Institute for Brain Science > > 551 N. 34th Street, Suite 200 > > Seattle, WA 98103 > > > > ralphpu@alleninstitute.org > > Tel: 206-548-7041 Fax: 206-548-7071 > > www.brainatlas.org > > > > _______________________________________________ > > Histonet mailing list > > Histonet@lists.utsouthwestern.edu > > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From histo <@t> compbio.com Tue Oct 18 08:36:21 2005 From: histo <@t> compbio.com (Histology) Date: Tue Oct 18 10:30:10 2005 Subject: [Histonet] position open Message-ID: <000201c5d3e8$ee036340$4075010a@steveyum> Dear Fellow Histonetters: We have a new position open for a histo tech-we are looking for a histotech (preferably) with good skills in animal tissues including grossing, embedding and sectioning. We have nice labs, great benefits, a favorable pay scale and are located in Sunnyvale, CA. We would also consider a trainee who wants to learn. Please respond to Dr. Poonam Kuruganti Poonam_kurganti@compbio.com 408 738 9260 who will give more details. cm From PMonfils <@t> Lifespan.org Tue Oct 18 10:34:24 2005 From: PMonfils <@t> Lifespan.org (Monfils, Paul) Date: Tue Oct 18 10:34:36 2005 Subject: [Histonet] Nissl Staining on Thick Sections Message-ID: <09C945920A6B654199F7A58A1D7D1FDE017175F8@lsexch.lsmaster.lifespan.org> Does anyone routinely do Nissl staining on 30 to 40 micron frozen sections? That's what the current project calls for, and I am finding that Nissl methods designed for 6 to 8 micron sections don't work well on the thick sections. They don't differentiate properly. If anyone has a Nissl protocol that works well on such thick sections, I would be grateful for any information. Paul M. From TJasper <@t> smdc.org Tue Oct 18 10:42:04 2005 From: TJasper <@t> smdc.org (Jasper, Thomas G.) Date: Tue Oct 18 10:42:44 2005 Subject: [Histonet] Block cutting protocols Message-ID: <1DB04B57B04C5747B87C3B39F3E605AA07D39094@harrier> Susan, First of all, the obvious answer to your question is a resounding NO. And (I believe) this is a cardinal rule which should not be broken. You state that you want "unbiased" responses, due to unknown circumstances by the members of the histonet. That's fine, but how in the world is there bias in this situation. It's cut and dried, you don't do it ever, under any circumstances. It's like lighting a cigarette while pumping gas! I do suspect that you may be trying to convince an administrator or some higher up with limited knowledge of clinical pathology of this fact. Or, you may be dealing with a "maverick" tech, that believes this enhances speed in sectioning or something along those lines. I may be wrong about all this, but whatever your reasons are good luck to you as this truly is a "no-brainer". tj -----Original Message----- From: Histology SLU [mailto:sluhisto@yahoo.com] Sent: Monday, October 17, 2005 9:05 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Block cutting protocols Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail communication and any attachments may contain confidential and privileged information for the use of the designated recipients named above. If you are not the intended recipient, you are hereby notified that you have received this communication in error and that any review, disclosure, dissemination, distribution or copying of it or its contents is prohibited. As required by federal and state laws, you need to hold this information as privileged and confidential. If you have received this communication in error, please notify the sender and destroy all copies of this communication and any attachments. From relia1 <@t> earthlink.net Tue Oct 18 11:28:11 2005 From: relia1 <@t> earthlink.net (Pam Barker) Date: Tue Oct 18 11:28:20 2005 Subject: [Histonet] RELIA Histology Job Opportunity Update 10/18/05 - additional openings from yesterday's posting!! Message-ID: Hello Histonetters, I hope you are enjoying a wonderful Autumn. Halloween is just around the corner TRICK or TREAT!!!!! The TRICK is finding the right job opportunity for you. And the TREAT is with my help it can be relatively painless. I will help you with your resume, coach you through the interview and offer process and refer you to positions based on the critieria you give me. Here are my current histology openings. All of the positions I work with are fulltime 40 hour per week positions with top hospitals, labs and doctors offices. My clients offer excellent compensation including competitive salaries, great benefits, relocation assistance and in some cases sign on bonuses. My services are FREE of charge to you. All of my fees are paid by my clients, (the facilities that I represent). I represent companies nationwide that are in need of histology supervisors, histotechnologists and histo technicians. Here is a list of my newest openings 1. Senior Research Specialist Immunohistochemistry ? California 2. Lead Tech (Clinical Setting) ? Dallas, Texas 3. Histo Tech (Clinical Setting) ? Dallas, Texas 4. Histo Tech (2 positions) Clinical Setting - SE Georgia 5. Histo Tech (2 positions) Clinical Setting - SW Florida (eligibility for FL lic req) 6. Technical Support Specialist - Research Setting - Northern California 7.Capital Equipment Sales ? Multiple U.S. locations 8. Histo Tech ? Clinical Setting (Northern CA/Southern OR) Here is a list of my current openings: HISTOLOGY MANAGEMENT: 1. Anatomic Pathology Manager ? Boston, MA 2. Histology Supervisor ? Boston, MA 3. Histology Supervisor ? South Florida HISTOTECHNICIAN/TECHNOLOGIST: 1. Histo Tech (clinical setting) ? Northern, CA 2. Histo Tech (research setting) ? Northern, CA 3. Histo Tech (clinical setting) ? Central Florida 4. Histo Tech (clinical setting) - Southeastern Florida 5. Histo Tech (clinical setting) ? Boston, MA 6. Histo Tech (clinical setting) - Minnesota 7. Histo Tech (clinical setting) ? Alaska If you or any of your friends would like more information on any of these positions. Or if you would like to discuss opportunities in other areas or future job searches please contact me. I would be more than happy to assist you. You can reach me at 866-607-3542 or relia1@earthlink.net I work with facilities nationwide and I will keep your resume confidential. I will only represent you to jobs you tell me you are interested in looking into. Remember It never hurts to keep an eye open even if you are happy in your present job. p.s. It was great to meet you at the NSH !!!!!!!!!! If our paths didn't cross at this year's NSH perhaps we will meet in Arizona!!! Happy Halloween!!!!!!!!!!!! Pam ? 866-607-3542 (866-60RELIA) Thank You! Pam Barker President RELIA Specialists in Allied Healthcare Recruiting 5703 Red Bug Lake Road #330 Winter Springs, FL 32708-4969 Phone: (407)657-2027 Cell: (407)353-5070 FAX: (407)678-2788 E-mail: relia1@earthlink.net From BDUE <@t> PARTNERS.ORG Tue Oct 18 11:31:52 2005 From: BDUE <@t> PARTNERS.ORG (Due, Brice) Date: Tue Oct 18 11:32:03 2005 Subject: [Histonet] RE: Nissl on Thick Paraffin... Message-ID: <59C772E2D8EDF345AE1601F5B60B3CB5010271A6@PHSXMB7.partners.org> Hello Nissl, I deleted your original post by accident & histosearch.com appears down... Try John Kiernan's modified cv nissl. It works more progressively than the other cv nissls I know, which are purely regressive. 100ml 0.1% cresyl violet spiked with the addition of 0.5ml of 1% oxalic acid. See p124 of his book. I do nissls on 15-20um paraffin, not the 40um you're talking. Even with John's oxalic nissl, I still like to overstain and regress with rosin gum. Gives a cleaner background = higher contrast. The rosin is a messy sticky sticky procedure, but you can control the differentiation by diluting the rosin. After aqueous cv staining of your choice, dehydrate slides in 95% etoh. Use 1-10% rosin gum in 95% etoh to differentiate. Make a 10% stock and cut when you need finer / slower control. You need a couple changes of rosin soln because it gets dirty fast. Dip slides in rosin until stain starts running, then rinse in 95% etoh and check on an old scope. Repeat until you get the contrast you want. If you take out too much cv, just re-hydrate and start over. The main problem with ultra-thick sections is that the front exposed side of the tissue will differentiate faster that the back side which is "hidden" against the slide. I would try diluting your differentiator (whatever procedure you're using) 1:10 and see if you can gain some control that way. Are free-floating sections an option? I've never tried that with paraffin embedded stuff. Or maybe try a non-cv based progressive nissl. I don't have a good recommendation there. Neutral red? Good Luck! -brice Neuropathology Lab Brigham & Women's Hospital Boston "THE INFORMATION TRANSMITTED IN THIS EMAIL IS INTENDED ONLY FOR THE PERSON OR ENTITY TO WHICH IT IS ADDRESSED AND MAY CONTAIN CONFIDENTIAL AND/OR PRIVILEGED MATERIAL. ANY REVIEW, RETRANSMISSION, DISSEMINATION OR OTHER USE OF OR TAKING OF ANY ACTION IN RELIANCE UPON, THIS INFORMATION BY PERSONS OR ENTITIES OTHER THAN THE INTENDED RECIPIENT IS PROHIBITED. IF YOU RECEIVE THIS EMAIL IN ERROR, PLEASE CONTACT THE SENDER AND DELETE THE MATERIAL FROM ANY COMPUTER." HIPPA Policy, 2003, Brigham & Women's Hospital, Boston, MA From fjones <@t> namsa.com Tue Oct 18 11:32:08 2005 From: fjones <@t> namsa.com (Fawn Jones) Date: Tue Oct 18 11:32:17 2005 Subject: [Histonet] Block cutting protocols Message-ID: <915E55B02E236E4D95258B181EEF6317F8FB9F@namsams01.namsa.int> I do believe that doing that would be a terrible idea, it leaves to many windows of opportunity for screw ups. I would never take the chance being that you could accidentally mix up your blocks or forget the order the sections went in. This would lead to many mistakes and possible misdiagnosis. It is an extremely bad idea Fawn Jones -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Jasper, Thomas G. Sent: Tuesday, October 18, 2005 11:42 AM To: 'Histology SLU' Cc: 'histonet@lists.utsouthwestern.edu' Subject: RE: [Histonet] Block cutting protocols Susan, First of all, the obvious answer to your question is a resounding NO. And (I believe) this is a cardinal rule which should not be broken. You state that you want "unbiased" responses, due to unknown circumstances by the members of the histonet. That's fine, but how in the world is there bias in this situation. It's cut and dried, you don't do it ever, under any circumstances. It's like lighting a cigarette while pumping gas! I do suspect that you may be trying to convince an administrator or some higher up with limited knowledge of clinical pathology of this fact. Or, you may be dealing with a "maverick" tech, that believes this enhances speed in sectioning or something along those lines. I may be wrong about all this, but whatever your reasons are good luck to you as this truly is a "no-brainer". tj -----Original Message----- From: Histology SLU [mailto:sluhisto@yahoo.com] Sent: Monday, October 17, 2005 9:05 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Block cutting protocols Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail communication and any attachments may contain confidential and privileged information for the use of the designated recipients named above. If you are not the intended recipient, you are hereby notified that you have received this communication in error and that any review, disclosure, dissemination, distribution or copying of it or its contents is prohibited. As required by federal and state laws, you need to hold this information as privileged and confidential. If you have received this communication in error, please notify the sender and destroy all copies of this communication and any attachments. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Melissa.Gonzalez <@t> cellgenesys.com Tue Oct 18 12:15:26 2005 From: Melissa.Gonzalez <@t> cellgenesys.com (Melissa Gonzalez) Date: Tue Oct 18 12:16:50 2005 Subject: [Histonet] RE: Nissl Staining on Thick Sections Message-ID: Paul, have you tried Molecular Probes? Their Nissl stains work great, you just have to adj. [ ]'s to your needs. Super fast and easy, and very bright specific results. Melissa ----------------------------- Message: 2 Date: Tue, 18 Oct 2005 11:34:24 -0400 From: "Monfils, Paul" Subject: [Histonet] Nissl Staining on Thick Sections To: "'histonet@lists.utsouthwestern.edu'" Message-ID: <09C945920A6B654199F7A58A1D7D1FDE017175F8@lsexch.lsmaster.lifespan.org> Content-Type: text/plain; charset="iso-8859-1" Does anyone routinely do Nissl staining on 30 to 40 micron frozen sections? That's what the current project calls for, and I am finding that Nissl methods designed for 6 to 8 micron sections don't work well on the thick sections. They don't differentiate properly. If anyone has a Nissl protocol that works well on such thick sections, I would be grateful for any information. Paul M. From pruegg <@t> ihctech.net Tue Oct 18 12:25:46 2005 From: pruegg <@t> ihctech.net (Patsy Ruegg) Date: Tue Oct 18 12:25:52 2005 Subject: [Histonet] mouse epithelial cells Message-ID: <200510181725.j9IHPeeu012317@chip.viawest.net> what antibody is being used to identify epithelial cells in mouse tissue. is everyone using a mouse cytokeratin with a MOM detection. are there any rabbit monoclonal cytokeratins out that work on ms tissue? Thanks, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. From ralphpu <@t> alleninstitute.org Tue Oct 18 12:25:59 2005 From: ralphpu <@t> alleninstitute.org (Ralph Puchalski) Date: Tue Oct 18 12:26:08 2005 Subject: [Histonet] Pink precipitate is monoformazan from BCIP/NBT reactiononISH? Message-ID: Dear John, Thank you for your detailed reply. In my earnestness to respond to your previous suggestions, and my desire to find a suitable blocking reagent for NBT, I forgot to mention that we have been up and running 4-5 robots per day for over a year, finishing over 10,000 genes in the mouse genome with a great number of positive (and negative) control probes that correspond exactly to the published in situ hybridization and microarray literature! We will be coming out with publications mining these data, and will have a presence at the Society for Neuroscience in November. Point number 3 is the most important. Could you recommend someone to us that could help with the NBT analogue idea? Maybe a biochemist or chemist? Number 4 also has a question for you. My responses to your 11 thoughts: 1. Yes, we had initially, and are having, success with the method you mentioned. 2. No, all reagent concentrations were initially chosen on the basis of RNA in situ hybridization experiments (tyramide-avidin-biotin-BCIP/NBT) using 25 um frozen sections with RNA probes designed according to published literature, and then compared with radio-labeled probes, which showed concurrence. Yes, the excessive monoformazan deposits are probably just a result of using concentrations of reagents and reaction times that were initially optimized for detection of the lowest levels of target mRNAs. We cannot monitor the colorimetric reaction for every probe, so we established an incubation period for all slides and all probes of 40 min in the BCIP/NBT/Levamisole buffer(Tris-HCl, NaCl, and MgCl2, Tween 20), which involves 2 x 15 min incubations and 1 x 10 min incubation, each incubation being a new aliquot of reagents. Certainly this is excessive for many probes, and leads to background issues including monoformazan, but it is standardized for all probes. The literature states that only in the ideal reaction does 2 moles of indoxyl fully reduce 1 mole of ditetetrazolium to produce 1 mole of insoluble diformazan. More often than not, the result is 2 moles of half-formazan (Smejkal and Kaul, JHC, 2001; original reference not available). 3. Yes, I am glad we agree on how NBT could bind to white matter preferentially since is it a lipophilic cation. My original question was how to block this binding. We already use several blocking steps (please see the web address I gave you) designed to inhibit non-specific binding of protein reagents, but no steps designed for the blocking of NBT to the tissue. Do you know of any analogues of NBT that are not substrates or inhibitors of alkaline phosphatase, are colorless, and would not be reduced by BCIP or a general reducing agent? 4. Yes to A and B. For C, we tried to wash excess or non-specifically bound substrates and products after the in situ hybridization (ISH) is finished, but water, acetone, or 95% ethanol without success (no decrease in ISH signal). We will try acid-alcohol washes soon. Just after inhibiting and washing off the completed colorimetric reaction, we treat the sections with 4% paraformaldehyde for 10 min. Do you think this step could serve to fix the non-specifically bound NBT and associated products to the tissue so washing them off after ISH becomes even more difficult? What kind of washes would you recommend? 5. Yes, please see my comment #2. 6. Agreed. 7. Okay. 8. Incorrect; see my reply #2. 9. Miscommunication/misunderstanding; see my reply #2. 10. Miscommunication/misunderstanding; see my reply #2. 11. Miscommunication/misunderstanding; see my reply #2. Thanks! Ralph -----Original Message----- From: John A. Kiernan [mailto:jkiernan@uwo.ca] Sent: Tuesday, October 18, 2005 8:28 AM To: Ralph Puchalski Cc: histonet@lists.utsouthwestern.edu; Johnson, Teri; Gayle Callis Subject: Re: [Histonet] Pink precipitate is monoformazan from BCIP/NBT reactiononISH? Dear Ralph, Eleven thoughts follow. 1. You state that "The incubation periods and concentrations were optimized for detection of the lowest levels of target RNA expression." Does this mean you initially had success with in situ hybridization, biotinylated tyramide amplification of the peroxidase label and further amplification by an avidin-alkaline phosphatase step and BCIP-nitroBT? 2. Were the riboprobe concentrations, times, temperatures etc chosen on the basis of some other method? If they were based on stained blots, especially if the hybridization signals were detected by a different method, the same conditions probably would not be optimal for sections of tissue. In a blot, the mRNA concentrations are very low because each band has an area of several square millimetres. In a section the local concentration in a cell that's busy expressing a gene is going to be much higher, with the expressing cell surrounded by other stuff (mostly parts of other cells , in brain tissue) that do not contain that mRNA. [Sorry for that very long sentence!] Your sections might be receiving too much riboprobe (nonspecific attachment and staining everywhere , albeit in the wrong colour. Alternatively the sections might be failing to bind any riboprobe, and the pink monoformazan is just a consequence of excessive incubation in the last stage of a ridiculously complicated method that has detected nothing. 3. NitroBT is a large lipophilic cation (Horobin 2002; Chapter 5, p.166 in the 10th edition of Conn's Biological Stains). As such, it can be expected to adhere more to white matter than to grey matter in either frozen or paraffin sections 4. When you deliberately reduced the nitroBT with ascorbate, in known hybridization-negative sections, you proved that (a) your alkaline phosphatase detection method could generate blue diformazan, (b) the natural disposition of the nitroBT was what you would expect from its well documented chemistry and histochemistry, and (c) you may not washing sections well enough to remove weakly bound reagents. 5. Have you seen localization of any well studied mRNA in a well documented site in the brain? If not, your procedure will never localize the sites of transcription or translation of any genes. 6. You state "First, a succession of blocking steps inhibits endogenous protein activity from interfering with ..." That's standard practice in immunohistochemistry and in situ hybridization, but the blocking reactions need ro be understood by the researcher. Some blocking methods can, if misused, provide wrong false-negative controls. 7. I haven't yet visited http://www.brain-map.org/pdf/ABADataProductionProcesses.pdf;jsessionid=9 FF15AEC860CC06CC8ECD7E42FF46402 to review the technical details of the methods carried out by your robots. I may have a look later in the week, but don't expect a detailed review by way of Histonet. 8. My guess is that your in situ hybridization technique, despite its multiple amplifications, is detecting nothing when applied to sections. 9. A possible explanation for the pink nothingness may be your "succession of blocking steps." Is there one that extracts mRNA or interferes with subsequent hybridization? Protein blocking procedures might prevent the oxidation products of biotinylated tyramine from binding to the tyrosine side-chains assumed to occur universally in animal tissues, 10. Before setting the robots to work on 20,000 genes, try to get the method to work under human control for 5 genes. If difficulties arise with even one of the five, there can be no justification for investing 4000 times as much money in applying the technology on a larger scale. 11. In every kind of histological or histochemical staining the chance of a fatal error increases with the number of steps in the technique and with the number of slides being processed. You need to get your method right for individual slides and then groups of 10, long before attempting to process 800 slides in a day. John Kiernan Anatomy, UWO London, Canada --------------------------- Ralph Puchalski wrote: > > Dear John, > > Thank you for your advice. Yes, we use digoxigenin (DIG)-labeled RNA > probes (riboprobes) in the in situ hybridization of frozen sections to > map gene expression throughout the mouse brain. Detection of bound > riboprobe is a multi-step procedure. First, a succession of blocking > steps inhibits endogenous protein activity from interfering with the > colorimetric enzymatic reactions. The colorimetric reaction itself is a > four part process, starting with the addition of a peroxidase-conjugated > antibody directed at the DIG-UTP hapten incorporated in the bound > riboprobe. A Tyramide Signal Amplification step is utilized to maximize > sensitivity. In brief, biotin-coupled tyramide is added to the tissue, > resulting in the formation of multiple activated tyramide molecules > through the activity of each bound peroxidase-coupled antibody molecule. > These highly reactive tyramide radicals bind to protein residues in the > vicinity of the bound riboprobe, thereby resulting in an amplification > of bound biotin molecules available for detection by up to a hundred > fold (relative to the number of bound antibody molecules). These biotin > molecules are then bound to NeutrAvidin-AP, the third step of the > colorimetric reaction. Alkaline phosphatase (AP) conjugated to the > NeutrAvidin (NA-AP) enzymatically cleaves the phosphate from > 5-bromo-4-chloro-3-indolyl phosphate (BCIP), and two of the resulting > indoles undergo a redox reaction with nitroblue tetrazolium (NBT) to > produce a blue particulate precipitate at the sites of riboprobe > binding. We process about 800 slides (1600 sections per day), and you > can visit our site for more details of our method and product at: > http://www.brain-map.org/pdf/ABADataProductionProcesses.pdf;jsessionid=9 > FF15AEC860CC06CC8ECD7E42FF46402 > > Processing 800 slides per day in order to cover the entire genome of > about 20,000 genes necessitates the use of robots. The slides are > incubated in only 300 ul of a particular reagent solution in chambers > mounted on an angle. The incubation periods and concentrations were > optimized for detection of the lowest levels of target RNA expression. > The conditions were established for the entire project and major changes > cannot be made. The system was set up so that all sections receive the > same amount of all reagents, including BCIP, NBT, and NA-AP. > > You commented on nothing dehydrogenases. Our experiments demonstrate > that removing NA-AP from the reaction, while keeping all other steps as > they are in the control, produces sections that are transparent, i.e. > without blue or purple staining under bright-field microscopy. If > anything, the sections have a very faint yellow tint, the same as what > we observe in the bottles just after we prepare the BCIP/NBT substrate > in buffer with Levamisole. However, if we incubate these slides in a > reducing agent such as 50 mM alkaline ascorbate for 5 minutes in pH 9.5 > in Tris-Magnesium-Saline, the slides paired to the ones that are > transparent become stained deep purple-blue (no NA-AP). Therefore, if > we are interpreting the data correctly, nothing dehydrogenases > contribute nothing that is detectable, and ascorbate reduces NBT to > diformazan. Do you agree? > > The troubling point is that the amount of staining visualized by > reduction with ascorbate matches the staining by the pink precipitate > (formed in the presence of NA-AP in our production runs and controls), > which presumably is monoformazan as determined spectrophotometrically. > The staining levels are highest in the fiber tracts, brainstem, and > hypothalamus, but are visible throughout the sections to various > extents. The reason why this is troubling is that the staining does not > require NA-AP. The NBT (we will test to be sure BCIP is not required) > just sticks to these areas, and later ends up on production slides > (NA-AP+) as monoformazan or pink precipitate, which is our hypothesis. > Do you agree with this assessment? > > How would you go about decreasing the staining of the fiber tracts, > brainstem, and hypothalamus, and the rest of the section with NBT? We > plan to titrate the NBT, but cannot reduce signal levels. Our fixation > and dehydration process does not remove all the fats in the tissue, so > it is quite possible that the hydrophobic NBT gets partitioned into the > lipid phase. Would you attempt to change the BCIP/NBT/Levamisole buffer > (Tris-HCl, NaCl, and MgCl2, Tween 20) used in the incubation with NA-AP > for color development? > > Do you know if anyone has tried to block the endogenous sites to which > NBT adheres? Without knowing how the binding occurs, it is difficult to > suggest options. How would you go about characterizing the binding of > NBT to the sections? I wish it were as easy as finding a convenient > analogue of NBT that is colorless, does not inhibit AP activity, is not > reduced, and binds to the same sites as NBT does, thereby blocking NBT > from binding to these sites. Any suggestions? > > Your advice is appreciated. > > Ralph Puchalski > > -----Original Message----- > From: John Kiernan [mailto:jkiernan@uwo.ca] > Sent: Monday, September 26, 2005 9:42 PM > To: Ralph Puchalski > Cc: histonet@lists.utsouthwestern.edu > Subject: Re: [Histonet] Pink precipitate is monoformazan from BCIP/NBT > reaction onISH? > > Dear Ralph, > > Your email is full of abbreviations and jargon. > Am I right in thinking it's a question about > localizing alkaline phosphatase activity by an > indoxyl-tetrazolium method? If so, please provide > a reference to the original publication and let us > all know if you followed it exactly or made any > changes. Part of your email suggests that the > alkaline phosphatase activity is not endogenous > but part of an amplification system used in > in situ hybridization. The pH optima of endogenous > and label alkaline phosphatases differ. > > The later paragraphs of your message indicate that > you may not fully understand the significance of > the mono- and diformazan products of reduction of > nitro-BT. Both colours result from reduction of > the tetrazolium salt, but you need controls to > prove that the reduction was by bromochloroindoxyl > and not by other reducing agents (such as diaphorases) > in the tissue. Non-enzymatic reduction of tetrazolium > salts by -SH has been a well known artifact {"nothing > dehydrogenase") for 40-45 years. > > John A. Kiernan > Anatomy Dept, UWO > London, Canada. > ___________________________________________________ > Ralph Puchalski wrote: > > > > I am trying to figure out the origin of the pink precipitate in the > > image I posted on http://www.histonet.org/site_images_frame.asp > > > > Please go to the image entitled: Pink Precipitate ver2.jpg. It is at > > the top of the list on 9/26/05, posted by Ralph Puchalski. To see the > > pink precipitate artifact, please open the image and set the scroll > bars > > on the bottom and right side of the image at their 1/2 way points. It > > is ugly! > > > > I think this precipitate is the aggregation of the monoformazan > > intermediate generated from NBT (after dephosphorylation of BCIP by > > alkaline phosphatase) that is not completely reduced to diformazan, > the > > insoluble dark blue or black precipitate that labels cells expressing > > target mRNAs in our in situ hybridization reactions. > > > > We don't know how the monoformazan adheres to the sections of tissue. > > It appears to be non-covalent due to the tendency of the monoformazan > to > > migrate under the coverslip in the aqueous mounting medium that has > yet > > to dry, and form clumps or aggregates or pools as seen in the picture. > > The monoformazan is soluble in ethanol so doesn't form pools or > > aggregates. But as soon as it is exposed to an aqueous medium, it > > precipitates. > > > > If we mount the post-ISH tissue sections with an organic based > mounting > > medium like UV-CureMount (Instrumedics), the monoformazan might cause > > the entire section to turn to a brown tint as seen on > > http://www.histonet.org/site_images_frame.asp > > > > Please go to the image Brown Tint 1.jpg. It is 4th image from the top > > on 9/26, posted by Ralph Puchalski. The lower image is mounted with > UV > > CureMount, and the upper was with aqueous Hydro Matrix. There is no > > pooling or precipitation of monoformazan with UV CureMount, but I > think > > it does cause the entire section to turn brown. > > > > Question: How do we eliminate this problem, which I believe is > > monoformazan? If we reduce it fully using ascorbate, the section > (later > > mounted with HyrdoMatrix) turns dark blue or purple, the same color as > > our ISH signal. We have tried washing off the monoformazan with 100% > > ethanol prior to coverslipping, but only small amounts are removed. > > 100% acetone also does not work effectively. > > > > Please let me know if you have any ideas that might help us eliminate > > this problem. > > > > Thank you, > > > > Ralph > > > > Ralph Puchalski, Ph.D. > > Manager, Process Engineering and Automation > > Allen Institute for Brain Science > > 551 N. 34th Street, Suite 200 > > Seattle, WA 98103 > > > > ralphpu@alleninstitute.org > > Tel: 206-548-7041 Fax: 206-548-7071 > > www.brainatlas.org > > > > _______________________________________________ > > Histonet mailing list > > Histonet@lists.utsouthwestern.edu > > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From pruegg <@t> ihctech.net Tue Oct 18 12:40:24 2005 From: pruegg <@t> ihctech.net (Patsy Ruegg) Date: Tue Oct 18 12:40:31 2005 Subject: [Histonet] DAKO keratin Message-ID: <200510181740.j9IHeHeu016588@chip.viawest.net> has anybody tried this pc rabbit anti-keratin Z0622 which uses cow keratin as the immunogen in mouse tissue? Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. From dpahisto <@t> yahoo.com Tue Oct 18 12:46:27 2005 From: dpahisto <@t> yahoo.com (Cindy DuBois) Date: Tue Oct 18 12:46:36 2005 Subject: [Histonet] PAS vs CAS Message-ID: <20051018174627.96514.qmail@web33407.mail.mud.yahoo.com> My lab manager recently read the article by Vinnie Della Speranza et. al. on fungus staining. He wanted us to try the Chromic Acid Schiff's (CAS) procedure and compare it to our PAS procedure. Although I get some staining with the CAS, only about half of the fungus is staining and it is very light. I used the following procedure: 1. Clear & Hydrate slides (standard protocol). 2. Incubate slides in 5% Chromic acid in water bath (temp approx. 42C) for 10minutes. Solution was pre-heated for 10 minutes. 3. Rinse and incubate in Schiff's solution for 10 minutes. 4. Rinse in hot water. (Our tap water has a lot of sulfur in it and therefore we do not normally do the sulfuric acid rinse). 5. Counterstain in Working Light Green, dehydrate and coverslip. If any one is using this procedure for their fungus stain would you please send me your procedure? Cindy DuBois, HT ASCP Integrated Pathology Associates Stockton CA --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From kl2215 <@t> columbia.edu Tue Oct 18 14:04:00 2005 From: kl2215 <@t> columbia.edu (Kim Lopez) Date: Tue Oct 18 14:04:10 2005 Subject: [Histonet] rat-specific nuclear ab or y-chromosome specific ab Message-ID: <1129662240.4355472069004@cubmail.cc.columbia.edu> has anybody heard of a rat-specific nuclear antibody or alternatively a y-chromosome specific antibody? thanks, Kim From PBugelsk <@t> CNTUS.JNJ.COM Tue Oct 18 14:43:39 2005 From: PBugelsk <@t> CNTUS.JNJ.COM (Bugelski, Peter [CNTUS]) Date: Tue Oct 18 14:44:01 2005 Subject: [Histonet] rat-specific nuclear ab or y-chromosome specific a b Message-ID: <7BF70FA941B9AE4783EAAF733762F1B5051981DC@CNTUSMAEXS3.na.jnj.com> Please include me on the reply list. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Kim Lopez Sent: Tuesday, October 18, 2005 3:04 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] rat-specific nuclear ab or y-chromosome specific ab has anybody heard of a rat-specific nuclear antibody or alternatively a y-chromosome specific antibody? thanks, Kim _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jkiernan <@t> uwo.ca Tue Oct 18 13:25:05 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Tue Oct 18 14:45:15 2005 Subject: [Histonet] RE: Nissl on Thick Paraffin... References: <59C772E2D8EDF345AE1601F5B60B3CB5010271A6@PHSXMB7.partners.org> Message-ID: <43553E01.B4CE5B1@uwo.ca> Cresyl violet with oxalic acid isn't one of mine! This is the first I've heard about it. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Due, Brice" wrote: > > Hello Nissl, I deleted your original post by accident & histosearch.com appears > down... > > Try John Kiernan's modified cv nissl. It works more progressively than the other > cv nissls I know, which are purely regressive. 100ml 0.1% cresyl violet spiked > with the addition of 0.5ml of 1% oxalic acid. See p124 of his book. > > I do nissls on 15-20um paraffin, not the 40um you're talking. Even with John's > oxalic nissl, I still like to overstain and regress with rosin gum. Gives a > cleaner background = higher contrast. The rosin is a messy sticky sticky > procedure, but you can control the differentiation by diluting the rosin. > > After aqueous cv staining of your choice, dehydrate slides in 95% etoh. Use > 1-10% rosin gum in 95% etoh to differentiate. Make a 10% stock and cut when you > need finer / slower control. You need a couple changes of rosin soln because it > gets dirty fast. Dip slides in rosin until stain starts running, then rinse in > 95% etoh and check on an old scope. Repeat until you get the contrast you want. > If you take out too much cv, just re-hydrate and start over. > > The main problem with ultra-thick sections is that the front exposed side of the > tissue will differentiate faster that the back side which is "hidden" against > the slide. I would try diluting your differentiator (whatever procedure you're > using) 1:10 and see if you can gain some control that way. Are free-floating > sections an option? I've never tried that with paraffin embedded stuff. Or maybe > try a non-cv based progressive nissl. I don't have a good recommendation there. > Neutral red? > > Good Luck! > -brice > Neuropathology Lab > Brigham & Women's Hospital > Boston > > "THE INFORMATION TRANSMITTED IN THIS EMAIL IS INTENDED ONLY FOR THE PERSON OR > ENTITY TO WHICH IT IS ADDRESSED AND MAY CONTAIN CONFIDENTIAL AND/OR PRIVILEGED > MATERIAL. ANY REVIEW, RETRANSMISSION, DISSEMINATION OR OTHER USE OF OR TAKING > OF ANY ACTION IN RELIANCE UPON, THIS INFORMATION BY PERSONS OR ENTITIES OTHER > THAN THE INTENDED RECIPIENT IS PROHIBITED. IF YOU RECEIVE THIS EMAIL IN ERROR, > PLEASE CONTACT THE SENDER AND DELETE THE MATERIAL FROM ANY COMPUTER." > HIPPA Policy, 2003, Brigham & Women's Hospital, Boston, MA > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From BDUE <@t> PARTNERS.ORG Tue Oct 18 15:05:43 2005 From: BDUE <@t> PARTNERS.ORG (Due, Brice) Date: Tue Oct 18 15:05:54 2005 Subject: [Histonet] RE: Nissl on Thick Paraffin... Message-ID: <59C772E2D8EDF345AE1601F5B60B3CB5010271A7@PHSXMB7.partners.org> John, I'm being overly broad, I'm sure, by calling it a "nissl". You list it as a counterstain for LFB, p124 of your text. Conceptually I always think of LFB+CV (=KB) as being LFB plus Nissl counterstain. The "KB" procedure here is exactly that, an LFB followed by a Nissl. Can a given cv procedure be a counterstain, but not a nissl when used as a primary stain? Sloppy terminology is very probably my mistake. My apologies. However your oxalic cv works well for me as a nissl and is cleaner than the purely regressive one on the books here. In any case, when I read another reply I realized the poster is working with FS, so I shouldn't have replied in the first place. Too little sleep, too much caffeine... Thanks, -brice "THE INFORMATION TRANSMITTED IN THIS EMAIL IS INTENDED ONLY FOR THE PERSON OR ENTITY TO WHICH IT IS ADDRESSED AND MAY CONTAIN CONFIDENTIAL AND/OR PRIVILEGED MATERIAL. ANY REVIEW, RETRANSMISSION, DISSEMINATION OR OTHER USE OF OR TAKING OF ANY ACTION IN RELIANCE UPON, THIS INFORMATION BY PERSONS OR ENTITIES OTHER THAN THE INTENDED RECIPIENT IS PROHIBITED. IF YOU RECEIVE THIS EMAIL IN ERROR, PLEASE CONTACT THE SENDER AND DELETE THE MATERIAL FROM ANY COMPUTER." HIPPA Policy, 2003, Brigham & Women's Hospital, Boston, MA -----Original Message----- From: John A. Kiernan [mailto:jkiernan@uwo.ca] Sent: Tuesday, October 18, 2005 2:25 PM To: Due, Brice Cc: histonet Subject: Re: [Histonet] RE: Nissl on Thick Paraffin... Cresyl violet with oxalic acid isn't one of mine! This is the first I've heard about it. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Due, Brice" wrote: > > Hello Nissl, I deleted your original post by accident & histosearch.com appears > down... > > Try John Kiernan's modified cv nissl. It works more progressively than the other > cv nissls I know, which are purely regressive. 100ml 0.1% cresyl violet spiked > with the addition of 0.5ml of 1% oxalic acid. See p124 of his book. > > I do nissls on 15-20um paraffin, not the 40um you're talking. Even with John's > oxalic nissl, I still like to overstain and regress with rosin gum. Gives a > cleaner background = higher contrast. The rosin is a messy sticky sticky > procedure, but you can control the differentiation by diluting the rosin. > > After aqueous cv staining of your choice, dehydrate slides in 95% etoh. Use > 1-10% rosin gum in 95% etoh to differentiate. Make a 10% stock and cut when you > need finer / slower control. You need a couple changes of rosin soln because it > gets dirty fast. Dip slides in rosin until stain starts running, then rinse in > 95% etoh and check on an old scope. Repeat until you get the contrast you want. > If you take out too much cv, just re-hydrate and start over. > > The main problem with ultra-thick sections is that the front exposed side of the > tissue will differentiate faster that the back side which is "hidden" against > the slide. I would try diluting your differentiator (whatever procedure you're > using) 1:10 and see if you can gain some control that way. Are free-floating > sections an option? I've never tried that with paraffin embedded stuff. Or maybe > try a non-cv based progressive nissl. I don't have a good recommendation there. > Neutral red? > > Good Luck! > -brice > Neuropathology Lab > Brigham & Women's Hospital > Boston > > "THE INFORMATION TRANSMITTED IN THIS EMAIL IS INTENDED ONLY FOR THE PERSON OR > ENTITY TO WHICH IT IS ADDRESSED AND MAY CONTAIN CONFIDENTIAL AND/OR PRIVILEGED > MATERIAL. ANY REVIEW, RETRANSMISSION, DISSEMINATION OR OTHER USE OF OR TAKING > OF ANY ACTION IN RELIANCE UPON, THIS INFORMATION BY PERSONS OR ENTITIES OTHER > THAN THE INTENDED RECIPIENT IS PROHIBITED. IF YOU RECEIVE THIS EMAIL IN ERROR, > PLEASE CONTACT THE SENDER AND DELETE THE MATERIAL FROM ANY COMPUTER." > HIPPA Policy, 2003, Brigham & Women's Hospital, Boston, MA > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From llewllew <@t> shaw.ca Tue Oct 18 15:05:58 2005 From: llewllew <@t> shaw.ca (Bryan Llewellyn) Date: Tue Oct 18 15:06:16 2005 Subject: [Histonet] PAS vs CAS References: <20051018174627.96514.qmail@web33407.mail.mud.yahoo.com> Message-ID: <001401c5d41f$5b4aa360$690a4246@yourlk4rlmsu> Try oxidising in the 5% chromic acid for about an hour. That was Bauer's original recommendation (4% for 40-60 minutes). Alternatively, many have found a 10% solution for 10 minutes satisfactory. You could also do a double oxidation with periodic acid. That is oxidise in 1% periodic acid for 15 minutes, block aldehyde with aniline-acetic for 30 minutes, then re-oxidise for 15 minutes. This also reduces background positive PAS staining. Bryan Llewellyn Bryan Llewellyn ----- Original Message ----- From: "Cindy DuBois" To: "Histonet" Sent: Tuesday, October 18, 2005 10:46 AM Subject: [Histonet] PAS vs CAS > My lab manager recently read the article by Vinnie Della Speranza et. al. > on fungus staining. He wanted us to try the Chromic Acid Schiff's (CAS) > procedure and compare it to our PAS procedure. > > Although I get some staining with the CAS, only about half of the fungus > is staining and it is very light. > > I used the following procedure: > 1. Clear & Hydrate slides (standard protocol). > 2. Incubate slides in 5% Chromic acid in water bath (temp approx. 42C) > for 10minutes. > Solution was pre-heated for 10 minutes. > 3. Rinse and incubate in Schiff's solution for 10 minutes. > 4. Rinse in hot water. > (Our tap water has a lot of sulfur in it and therefore we do not normally > do the sulfuric acid rinse). > 5. Counterstain in Working Light Green, dehydrate and coverslip. > > If any one is using this procedure for their fungus stain would you please > send me your procedure? > > Cindy DuBois, HT ASCP > Integrated Pathology Associates > Stockton CA > > > > > > --------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > From llewllew <@t> shaw.ca Tue Oct 18 15:10:27 2005 From: llewllew <@t> shaw.ca (Bryan Llewellyn) Date: Tue Oct 18 15:11:09 2005 Subject: [Histonet] Nissl staining on thick sections Message-ID: <002801c5d41f$fb2be600$690a4246@yourlk4rlmsu> I also deleted the original question, but Nissl can also be stained progressively with an Unna-Pappenheim (methyl green-pyronin). For that purpose it is not necessary to remove methyl violet with chloroform extraction, and the simplest of methods usually works quite well. Of course, the Nissl is red instead of blue. Bryan Llewellyn From llewllew <@t> shaw.ca Tue Oct 18 15:14:01 2005 From: llewllew <@t> shaw.ca (Bryan Llewellyn) Date: Tue Oct 18 15:14:18 2005 Subject: [Histonet] PAS vs CAS References: <20051018174627.96514.qmail@web33407.mail.mud.yahoo.com> <001401c5d41f$5b4aa360$690a4246@yourlk4rlmsu> Message-ID: <003501c5d420$7b16c920$690a4246@yourlk4rlmsu> In my haste I didn't note the 42C temperature. Try it at room temperature. Bryan Llewellyn ----- Original Message ----- From: "Bryan Llewellyn" To: "Histonet" Sent: Tuesday, October 18, 2005 1:05 PM Subject: Re: [Histonet] PAS vs CAS > Try oxidising in the 5% chromic acid for about an hour. That was Bauer's > original recommendation (4% for 40-60 minutes). Alternatively, many have > found a 10% solution for 10 minutes satisfactory. You could also do a > double oxidation with periodic acid. That is oxidise in 1% periodic acid > for 15 minutes, block aldehyde with aniline-acetic for 30 minutes, then > re-oxidise for 15 minutes. This also reduces background positive PAS > staining. > > Bryan Llewellyn > > Bryan Llewellyn > ----- Original Message ----- > From: "Cindy DuBois" > To: "Histonet" > Sent: Tuesday, October 18, 2005 10:46 AM > Subject: [Histonet] PAS vs CAS > > >> My lab manager recently read the article by Vinnie Della Speranza et. al. >> on fungus staining. He wanted us to try the Chromic Acid Schiff's (CAS) >> procedure and compare it to our PAS procedure. >> >> Although I get some staining with the CAS, only about half of the fungus >> is staining and it is very light. >> >> I used the following procedure: >> 1. Clear & Hydrate slides (standard protocol). >> 2. Incubate slides in 5% Chromic acid in water bath (temp approx. 42C) >> for 10minutes. >> Solution was pre-heated for 10 minutes. >> 3. Rinse and incubate in Schiff's solution for 10 minutes. >> 4. Rinse in hot water. >> (Our tap water has a lot of sulfur in it and therefore we do not normally >> do the sulfuric acid rinse). >> 5. Counterstain in Working Light Green, dehydrate and coverslip. >> >> If any one is using this procedure for their fungus stain would you >> please send me your procedure? >> >> Cindy DuBois, HT ASCP >> Integrated Pathology Associates >> Stockton CA >> >> >> >> >> >> --------------------------------- >> Yahoo! Music Unlimited - Access over 1 million songs. Try it free. >> _______________________________________________ >> Histonet mailing list >> Histonet@lists.utsouthwestern.edu >> http://lists.utsouthwestern.edu/mailman/listinfo/histonet >> > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > From katherine-walters <@t> uiowa.edu Tue Oct 18 15:15:36 2005 From: katherine-walters <@t> uiowa.edu (Walters, Katherine S) Date: Tue Oct 18 15:15:43 2005 Subject: [Histonet] alcian blue and alizarin red Message-ID: Has anyone done alcian blue/alizarin red staining on tissue sections? The protocols I've turned up seem to be for cleared tissue. Thanks, Kathy From rjbuesa <@t> yahoo.com Tue Oct 18 15:32:36 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Tue Oct 18 15:32:50 2005 Subject: [Histonet] alcian blue and alizarin red In-Reply-To: Message-ID: <20051018203236.7059.qmail@web61225.mail.yahoo.com> Alizarin red was the standard procedure I used for calcium (carbonate / phosphates) pretty much as descibed by Sheehan & Hrapchak [(1973):153-4] The solution has to have pH adjusted to 4.1-4.3 and final dehydrating has to be done with acetone (as ethanol will remove the stain). Alcian blue is standard for mucosubstances [Luna,1968:163] but I never used both combined. Both procedures on sections of paraffin embedded tisusues. Rene J "Walters, Katherine S" wrote: Has anyone done alcian blue/alizarin red staining on tissue sections? The protocols I've turned up seem to be for cleared tissue. Thanks, Kathy _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From histo20 <@t> hotmail.com Tue Oct 18 18:12:52 2005 From: histo20 <@t> hotmail.com (Paula Wilder) Date: Tue Oct 18 18:13:01 2005 Subject: [Histonet] Ventana immunostainer Message-ID: Our institution is trying the new Ventana immunostainer. We have had the unit since July and everything was staining nicely. Then we started to get variable staining, especially with CK20, ER and PR, ki67. Ventana reps have been working diligently to get this problem resolved. We are using pre-dilutes and the staining seems to be fine the first time, then increasingly variable. The reps took our controls to another area hospital and tested them there and the staining was fine; hence, the problem is with our machine. I know they will replace this instrument if it continues to fail, but I was wondering if anyone out there has similar problems. Thanks for any help! Paula Wilder St. Joseph Medical Center 7601 Osler Drive Towson, MD 21204 410-337-1741 From lpwenk <@t> sbcglobal.net Tue Oct 18 20:15:28 2005 From: lpwenk <@t> sbcglobal.net (Lee & Peggy Wenk) Date: Tue Oct 18 20:16:05 2005 Subject: [Histonet] PAS vs CAS In-Reply-To: <003501c5d420$7b16c920$690a4246@yourlk4rlmsu> Message-ID: I think what Bryan suggested would work. For our GMS, we usually use 10% chromic acid for 10 minutes at room temp, and they look great. However, when I tried the 10%/10 min for the CAS, only about half the fungi stained. When I reduced the time to 5 minutes, more stained, and brighter pink. So I think the fungi were being over oxidized. So 5% at room temp for 10 minutes sounds like it should improve the staining. Let us know. Peggy Wenk, HTL(ASCP)SLS William Beaumont Hospital Royal Oak, MI 48073 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Bryan Llewellyn Sent: Tuesday, October 18, 2005 4:14 PM To: Histonet Subject: Re: [Histonet] PAS vs CAS In my haste I didn't note the 42C temperature. Try it at room temperature. Bryan Llewellyn ----- Original Message ----- From: "Bryan Llewellyn" To: "Histonet" Sent: Tuesday, October 18, 2005 1:05 PM Subject: Re: [Histonet] PAS vs CAS > Try oxidising in the 5% chromic acid for about an hour. That was Bauer's > original recommendation (4% for 40-60 minutes). Alternatively, many have > found a 10% solution for 10 minutes satisfactory. You could also do a > double oxidation with periodic acid. That is oxidise in 1% periodic acid > for 15 minutes, block aldehyde with aniline-acetic for 30 minutes, then > re-oxidise for 15 minutes. This also reduces background positive PAS > staining. > > Bryan Llewellyn > > Bryan Llewellyn > ----- Original Message ----- > From: "Cindy DuBois" > To: "Histonet" > Sent: Tuesday, October 18, 2005 10:46 AM > Subject: [Histonet] PAS vs CAS > > >> My lab manager recently read the article by Vinnie Della Speranza et. al. >> on fungus staining. He wanted us to try the Chromic Acid Schiff's (CAS) >> procedure and compare it to our PAS procedure. >> >> Although I get some staining with the CAS, only about half of the fungus >> is staining and it is very light. >> >> I used the following procedure: >> 1. Clear & Hydrate slides (standard protocol). >> 2. Incubate slides in 5% Chromic acid in water bath (temp approx. 42C) >> for 10minutes. >> Solution was pre-heated for 10 minutes. >> 3. Rinse and incubate in Schiff's solution for 10 minutes. >> 4. Rinse in hot water. >> (Our tap water has a lot of sulfur in it and therefore we do not normally >> do the sulfuric acid rinse). >> 5. Counterstain in Working Light Green, dehydrate and coverslip. >> >> If any one is using this procedure for their fungus stain would you >> please send me your procedure? >> >> Cindy DuBois, HT ASCP >> Integrated Pathology Associates >> Stockton CA >> >> >> >> >> >> --------------------------------- >> Yahoo! Music Unlimited - Access over 1 million songs. Try it free. >> _______________________________________________ >> Histonet mailing list >> Histonet@lists.utsouthwestern.edu >> http://lists.utsouthwestern.edu/mailman/listinfo/histonet >> > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jmahoney <@t> alegent.org Tue Oct 18 20:16:33 2005 From: jmahoney <@t> alegent.org (Janice A Mahoney) Date: Tue Oct 18 20:17:06 2005 Subject: [Histonet] Ventana immunostainer Message-ID: What kind of water are you using to make your solutions? Jan Omaha >>> "Paula Wilder" 10/18/2005 6:12 PM >>> Our institution is trying the new Ventana immunostainer. We have had the unit since July and everything was staining nicely. Then we started to get variable staining, especially with CK20, ER and PR, ki67. Ventana reps have been working diligently to get this problem resolved. We are using pre-dilutes and the staining seems to be fine the first time, then increasingly variable. The reps took our controls to another area hospital and tested them there and the staining was fine; hence, the problem is with our machine. I know they will replace this instrument if it continues to fail, but I was wondering if anyone out there has similar problems. Thanks for any help! Paula Wilder St. Joseph Medical Center 7601 Osler Drive Towson, MD 21204 410-337-1741 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From CDWallace <@t> wyeth.com Wed Oct 19 07:23:51 2005 From: CDWallace <@t> wyeth.com (Catherine D. Wallace) Date: Wed Oct 19 07:24:20 2005 Subject: [Histonet] Please take me off the histo list. Thank you. Message-ID: Please take me off the histo list. Thank you. From ROrr <@t> enh.org Wed Oct 19 08:09:16 2005 From: ROrr <@t> enh.org (Orr, Rebecca) Date: Wed Oct 19 08:09:26 2005 Subject: [Histonet] block cutting protocols Message-ID: Our IHC lab has 3 techs and our Histo Lab has 12 Techs....so we have a fairly large number to compare and NO ONE in our facility does this! As much as I have never seen any CAP regulation, I would not permit the practice of running more than one block of ribbons on the water bath... I believe that is a fundamental practice taught from the beginning of a Histo-tech's training. Besides the opportunity to pick up the wrong slide, there is contamination... the surface of the water bath should be inspected and/or "swiped" to rid of any previous particles after each ribbon.... You'd need to decide what is riskier...saving some time or sending a slide to a doc that has a part of another case on it. The answer is obvious. Becky Orr CLA,HT(ASCP) IHC Lead Evanston Northwestern Healthcare 847-570-2771 -----Original Message----- From: Histology SLU [mailto:sluhisto@yahoo.com] Sent: Monday, October 17, 2005 9:05 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Block cutting protocols Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan From jmahoney <@t> alegent.org Wed Oct 19 09:32:08 2005 From: jmahoney <@t> alegent.org (Janice A Mahoney) Date: Wed Oct 19 09:32:45 2005 Subject: Fwd: RE: [Histonet] Ventana immunostainer Message-ID: >>> "Cazares, Ruth" 10/19/2005 9:30 AM >>> Janice, I had a similar problem when we first got our Ventana instrument. What it turned out to be the slides. We ordered from a cheaper source and got spotty staining. We switched to the Fisher (+) slides and have not had a problem for the last 8 months. Good Luck! Ruth Cazares Swedish Covenant Hospital Chicago, IL -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Janice A Mahoney Sent: Tuesday, October 18, 2005 8:17 PM To: histo20@hotmail.com; histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Ventana immunostainer What kind of water are you using to make your solutions? Jan Omaha >>> "Paula Wilder" 10/18/2005 6:12 PM >>> Our institution is trying the new Ventana immunostainer. We have had the unit since July and everything was staining nicely. Then we started to get variable staining, especially with CK20, ER and PR, ki67. Ventana reps have been working diligently to get this problem resolved. We are using pre-dilutes and the staining seems to be fine the first time, then increasingly variable. The reps took our controls to another area hospital and tested them there and the staining was fine; hence, the problem is with our machine. I know they will replace this instrument if it continues to fail, but I was wondering if anyone out there has similar problems. Thanks for any help! Paula Wilder St. Joseph Medical Center 7601 Osler Drive Towson, MD 21204 410-337-1741 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet *** Confidentiality Statement *** This e-mail is intended only for the use of the individual or entity to which it is addressed and may contain information that is privileged and confidential. If the reader of this message is not the intended recipient, please notify the sender immediately by replying to this message and then delete it from your system. Any review, dissemination, distribution, or reproduction of this message by unintended recipients is strictly prohibited and may be subject to legal restriction. Thank you for your cooperation. From jmahoney <@t> alegent.org Wed Oct 19 10:33:22 2005 From: jmahoney <@t> alegent.org (Janice A Mahoney) Date: Wed Oct 19 10:34:00 2005 Subject: Fwd: RE: [Histonet] Ventana immunostainer Message-ID: Hello, I use Ventana benchmarks and the XT in my lab and have absolutely no problems. We have beautiful reproducible staining with every antibody we have worked up. Paula is the person who is having trouble with some antibodies. I think it is very important when bring up a new instrument or system that you eliminate any variables you can. I'm sure Ventana will give you some references of happy customers. Find out what a lab who has good consistent results uses as far a water, slides, fixation times, ptotocols etc. Most problems can be overcome from that point on with a little tweaking of diluents, cell conditioning, etc. Eliminating the variables on your controls is also important. Once you get antibodies up and running I think you will be very happy with Ventana's quality and reproducibility. Have your rep check the instrument for any instrument problems like pads not heating properly. Good Luck Jan Mahoney Omaha >>> "Janice A Mahoney" 10/19/2005 9:32 AM >>> >>> "Cazares, Ruth" 10/19/2005 9:30 AM >>> Janice, I had a similar problem when we first got our Ventana instrument. What it turned out to be the slides. We ordered from a cheaper source and got spotty staining. We switched to the Fisher (+) slides and have not had a problem for the last 8 months. Good Luck! Ruth Cazares Swedish Covenant Hospital Chicago, IL -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Janice A Mahoney Sent: Tuesday, October 18, 2005 8:17 PM To: histo20@hotmail.com; histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Ventana immunostainer What kind of water are you using to make your solutions? Jan Omaha >>> "Paula Wilder" 10/18/2005 6:12 PM >>> Our institution is trying the new Ventana immunostainer. We have had the unit since July and everything was staining nicely. Then we started to get variable staining, especially with CK20, ER and PR, ki67. Ventana reps have been working diligently to get this problem resolved. We are using pre-dilutes and the staining seems to be fine the first time, then increasingly variable. The reps took our controls to another area hospital and tested them there and the staining was fine; hence, the problem is with our machine. I know they will replace this instrument if it continues to fail, but I was wondering if anyone out there has similar problems. Thanks for any help! Paula Wilder St. Joseph Medical Center 7601 Osler Drive Towson, MD 21204 410-337-1741 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet *** Confidentiality Statement *** This e-mail is intended only for the use of the individual or entity to which it is addressed and may contain information that is privileged and confidential. If the reader of this message is not the intended recipient, please notify the sender immediately by replying to this message and then delete it from your system. Any review, dissemination, distribution, or reproduction of this message by unintended recipients is strictly prohibited and may be subject to legal restriction. Thank you for your cooperation. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From CDWallace <@t> wyeth.com Wed Oct 19 11:30:35 2005 From: CDWallace <@t> wyeth.com (Catherine D. Wallace) Date: Wed Oct 19 11:33:01 2005 Subject: [Histonet] Unscribe to Histonet Message-ID: Please take me off the histonet. Thank you.........Katy Wallace From clarissabush <@t> sbcglobal.net Wed Oct 19 11:35:50 2005 From: clarissabush <@t> sbcglobal.net (CM Bush) Date: Wed Oct 19 11:35:59 2005 Subject: [Histonet] How far will stain reach into a thick section on a slide? Message-ID: <20051019163550.17477.qmail@web80304.mail.yahoo.com> Dear Histonet, I am wondering what you can tell me. We are staining thick sections of human brain. The brain has been very well fixed in 10% NBF, and sections are mounted on slides. We are using thick sections (50um) and Nissl or immuno staining. Does any one know how deeply Nissl stain (or an IHC stain) can permeate tissue mounted on a slide? Is, say, any tissue thickness greater than 20um (for example) moot as it will never be reached by the stain? Would staining floating sections be a more rational way to deal with thick sections? Thank you vey much for the help. Clarissa From mward <@t> wfubmc.edu Wed Oct 19 14:21:16 2005 From: mward <@t> wfubmc.edu (Martha Ward) Date: Wed Oct 19 14:34:26 2005 Subject: [Histonet] Varicella zoster immuno controls Message-ID: <61135F0455D33347B5AAE209B903A304106A0ED4@EXCHVS2.medctr.ad.wfubmc.edu> I am looking for a vendor or someone out there who could share a positive VZV immuno control block. Thanks in advance for your help. Martha Ward Wake Forest University Baptist Medical Center From ewrona <@t> yahoo.com Wed Oct 19 17:13:34 2005 From: ewrona <@t> yahoo.com (Erin Wrona) Date: Wed Oct 19 17:13:44 2005 Subject: [Histonet] Looking for a job Message-ID: <20051019221335.22111.qmail@web52502.mail.yahoo.com> Hi all, Does anyone know of any job openings in Buffalo NY? Thanks, Erin From jkiernan <@t> uwo.ca Wed Oct 19 23:58:06 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Wed Oct 19 23:58:17 2005 Subject: [Histonet] How far will stain reach into a thick section on a slide? References: <20051019163550.17477.qmail@web80304.mail.yahoo.com> Message-ID: <435723DE.6A0C99DE@uwo.ca> Sections of brain much thicker than 20um can be Nissl stained or immunostained all the way through. For some purposes 80-100um is the preferred thickness. Nuclei (meaning groups of similar neurons) stand out clearly, and individual immunostained axons or long dendrites can often be traced over long distances, especially if there are not too many, close together. Usually thick frozen sections are handled free-floating, so that reagents can enter from both surfaces. If your 50um sections are stuck on slides, longer times may be needed. For immunostaining it's usual to apply the primary antibody for 24-48 hours at 4C and to include a non-ionic detergent such as saponin or Triton-X to make holes in the cell membranes. Times in secondary antisera and related reagents are only 30-60 minutes. For Nissl staining use a weak solution of a cationic dye at pH between 3.5 and 4.0). Stain progressively and wash in water buffered to the same or a higher pH. An acetate buffer is convenient and cheap. It can be very dilute but must not be even slightly more acidic than the dye solution. If you wash the sections in tap water, do this is in the sink, with a rapid flow of water through the slides. If the sections spend significant time in an unbuffered solution of the cationic dye there will be some background (protein) staining . A pure Nissl stain shows only DNA (cell nuclei) and RNA (in rough endoplasmic reticulum of neuronal cell bodies). --- Historical interlude: Franz Nissl (1860-1919) was a German "neuropsychiatrist" who also did the histopathology of his patients. He described normal and pathological neurons about 10 years before the biochemical recognition of nucleic acids and at least 50 years before cytoplasmic basophilia was solidly attributed to to RNA in ribosomes. If you find this sort of historical stuff interesting, click on http://www.whonamedit.com/doctor.cfm/2465.html for Nissl, the man and for links about his mentor, poor old Johan Bernhard Aloys von Gudden (1824-1886), who was probably murdered by an ungrateful patient, King Ludwig II of Bavaria. The mad king and the neuropsychiatrist died together. von Gudden had been sent by the parliament to certify Ludwig, who had brought his country close to bankruptcy by building fairytale castles and palaces. The buildings are still there, and tourism may have recovered the losses incurred by Ludwig II. --- Which dye? Thionine, toluidine blue and azure A are good blue Nissl stains; neutral red is also good. Many people like cresyl violet, and there's a chemically related dye called Darrow red that was invented for Nissl staining (MM Powers et al. 1960 Stain Technol 30:83-88). I've never used Darrow red or seen it offered for sale, but its use appeared in publications in the 1990s. Darrow red has also been used by biochemists as an enzyme inhibitor. The dye is named for the late Mary Darrow, who was the chief technician at the Biological Stain Commission's laboratory in Rochester, NY in the 1930s, '40s and '50s. Having found a suitable concentration and time for staining your sections in your lab, rinse in the acetate buffer and dehydrate in 3 changes of 100% alcohol. This will minimize loss of dye from the stained nucleic acids. Alcohol-water mixtures extract most stains more quickly than 100% alcohol. Several Nissl procedures exploit this, and use a graded alcohol dehydration to differentiate overstained sections. That's a waste of good alcohol. The specificity of staining is determined largely by the pH of the dye solution. The staining time will be shorter with a higher dye concentration, but for thick sections time is needed for penetration. If the dye concentration is too high there will be more unbound dye to wash out of the sections, and this will end up in the dehydrating alcohols. Go for the lowest dye concentration that fits into your working day. John Kiernan Anatomy, UWO London, Canada ----------------------------------------- CM Bush wrote: > > Dear Histonet, > > I am wondering what you can tell me. We are staining thick sections of human brain. The brain has been very well fixed in 10% NBF, and sections are mounted on slides. We are using thick sections (50um) and Nissl or immuno staining. Does any one know how deeply Nissl stain (or an IHC stain) can permeate tissue mounted on a slide? Is, say, any tissue thickness greater than 20um (for example) moot as it will never be reached by the stain? > Would staining floating sections be a more rational way to deal with thick sections? > > Thank you vey much for the help. > Clarissa > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From LJApuzzio <@t> msn.com Thu Oct 20 04:11:02 2005 From: LJApuzzio <@t> msn.com (Louis Apuzzio) Date: Thu Oct 20 04:11:16 2005 Subject: [Histonet] Histology Opportunity Message-ID: Hello Histonetters; I am asking assistance if any one knows of any available opportunities in Histology. I am HT eligible and also a CT(ASCP). It has been difficult for me finding an available opportunity. I have excellent skills and just need an open door. Thank you all very much. Have a great day. Louis J. Apuzzio From Susan.Ferrigon <@t> sanofi-aventis.com Thu Oct 20 08:35:40 2005 From: Susan.Ferrigon <@t> sanofi-aventis.com (Susan.Ferrigon@sanofi-aventis.com) Date: Thu Oct 20 08:35:55 2005 Subject: [Histonet] Mechanical tool for removing spinal cord from rats. Message-ID: Hi What do people use to remove spinal cords from rats during post mortems?? At present we use large scissors to cut through the bone but are looking to try a mechanical tool to replace this. Does any one have any recomendations.?? Thanks Susan ---------------------------------------------------------- Le pr?sent message ainsi que ses ?ventuelles pi?ces jointes est exclusivement destin? au(x) destinataire(s), personnes physiques ou morales, qu'il d?signe. Il constitue de ce fait une correspondance ? caract?re priv? et peut contenir des informations confidentielles. Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser imm?diatement l'exp?diteur par retour de courrier ?lectronique puis de le d?truire, ainsi que ses ?ventuelles pi?ces jointes, sans en conserver de copie. This message, including any attachment, is intended for the use of the individual or entity to which it is addressed. It is therefore to be considered as a private correspondence which may contain confidential information. If you are not the intended recipient, please advise the sender immediately by reply e.mail and delete this message and any attachment thereto without retaining a copy. ---------------------------------------------------------- From ree3 <@t> leicester.ac.uk Thu Oct 20 08:47:34 2005 From: ree3 <@t> leicester.ac.uk (Edwards, R.E.) Date: Thu Oct 20 08:47:46 2005 Subject: [Histonet] Mechanical tool for removing spinal cord from rats. Message-ID: Spinal Tap?. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Susan.Ferrigon@sanofi-aventis.com Sent: 20 October 2005 14:36 To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Mechanical tool for removing spinal cord from rats. Hi What do people use to remove spinal cords from rats during post mortems?? At present we use large scissors to cut through the bone but are looking to try a mechanical tool to replace this. Does any one have any recomendations.?? Thanks Susan ---------------------------------------------------------- Le pr?sent message ainsi que ses ?ventuelles pi?ces jointes est exclusivement destin? au(x) destinataire(s), personnes physiques ou morales, qu'il d?signe. Il constitue de ce fait une correspondance ? caract?re priv? et peut contenir des informations confidentielles. Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser imm?diatement l'exp?diteur par retour de courrier ?lectronique puis de le d?truire, ainsi que ses ?ventuelles pi?ces jointes, sans en conserver de copie. This message, including any attachment, is intended for the use of the individual or entity to which it is addressed. It is therefore to be considered as a private correspondence which may contain confidential information. If you are not the intended recipient, please advise the sender immediately by reply e.mail and delete this message and any attachment thereto without retaining a copy. ---------------------------------------------------------- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From pmarcum <@t> vet.upenn.edu Thu Oct 20 09:05:03 2005 From: pmarcum <@t> vet.upenn.edu (Pamela Marcum) Date: Thu Oct 20 09:05:16 2005 Subject: [Histonet] Mechanical tool for removing spinal cord from rats. In-Reply-To: References: Message-ID: <6.1.1.1.2.20051020094900.01997de8@mail.vet.upenn.edu> Hi Susan, I used a small rongeur that I found with the orthopedic hand surgeons. It had very small jaws that allowed me to go under the vertebrae and cleanly brake them away without injuring the spinal column. I just had to ask around in the surgical supply books and find one I liked. It looked like cuticle clippers manicurist used only the ends were rounded with a small cup like area. Let me know if this not a good enough description. I would remove the spinal cord intact with the brain through the cauda equina without damaging the cord. This is also allowed me to count down the ganglia and vertebrae as I went. Pam Marcum UPenn Vet School New Bolton Center 610-925-6278 At 09:35 AM 10/20/2005, Susan.Ferrigon@sanofi-aventis.com wrote: >Hi > >What do people use to remove spinal cords from rats during post mortems?? >At present we use large scissors to cut through the bone but are looking to >try a mechanical tool to replace this. >Does any one have any recomendations.?? > >Thanks > >Susan > >---------------------------------------------------------- >Le pr?sent message ainsi que ses ?ventuelles pi?ces jointes est >exclusivement destin? au(x) destinataire(s), personnes physiques ou >morales, qu'il d?signe. >Il constitue de ce fait une correspondance ? caract?re priv? et peut >contenir des informations confidentielles. >Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser >imm?diatement l'exp?diteur par retour de courrier ?lectronique puis de le >d?truire, ainsi que ses ?ventuelles pi?ces jointes, sans en conserver de >copie. > > >This message, including any attachment, is intended for the use of the >individual or entity to which it is addressed. >It is therefore to be considered as a private correspondence which may >contain confidential information. >If you are not the intended recipient, please advise the sender immediately >by reply e.mail and delete this message and any attachment thereto without >retaining a copy. >---------------------------------------------------------- > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Susan.Ferrigon <@t> sanofi-aventis.com Thu Oct 20 09:19:03 2005 From: Susan.Ferrigon <@t> sanofi-aventis.com (Susan.Ferrigon@sanofi-aventis.com) Date: Thu Oct 20 09:19:17 2005 Subject: [Histonet] To clarify : Mechanical tool for removing spinal cord from rats. Message-ID: Hi What do people use to remove spinal cords from rats during post mortems?? At present we use large scissors to cut through the bone but are looking to try a mechanical tool to replace this. Does any one have any recomendations.?? Thanks Susan Thanks to all who replied so quickly, To clarify, I am actually wanting to remove the whole spinal column with the cord inside, so it would be a tool to cut through the ribs attached to the spinal column, we do a lot of pm's and are trying to make life a little easier on the hands as cutting through the bone can be tough. Thanks once again Susan ---------------------------------------------------------- Le pr?sent message ainsi que ses ?ventuelles pi?ces jointes est exclusivement destin? au(x) destinataire(s), personnes physiques ou morales, qu'il d?signe. Il constitue de ce fait une correspondance ? caract?re priv? et peut contenir des informations confidentielles. Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser imm?diatement l'exp?diteur par retour de courrier ?lectronique puis de le d?truire, ainsi que ses ?ventuelles pi?ces jointes, sans en conserver de copie. This message, including any attachment, is intended for the use of the individual or entity to which it is addressed. It is therefore to be considered as a private correspondence which may contain confidential information. If you are not the intended recipient, please advise the sender immediately by reply e.mail and delete this message and any attachment thereto without retaining a copy. ---------------------------------------------------------- From mucram11 <@t> comcast.net Thu Oct 20 09:26:17 2005 From: mucram11 <@t> comcast.net (Pam Marcum) Date: Thu Oct 20 09:26:35 2005 Subject: [Histonet] To clarify : Mechanical tool for removing spinal cord from rats. Message-ID: <102020051426.21889.4357A90900067948000055812202888744CECE030E9D0C9A03@comcast.net> I would still look in the surgical instrument supply books for something like a rongeur that allows the user to control the cut of ribs and bone better with less trauma to the actual spinal coloumn and limit the movement during cutting. The cutting area of the rongeur is designed to cut bone cleanly and quickly. Hope this helps, Pam Marcum > > > > > Hi > > What do people use to remove spinal cords from rats during post mortems?? > At present we use large scissors to cut through the bone but are looking to > try a mechanical tool to replace this. > Does any one have any recomendations.?? > > Thanks > > Susan > > > Thanks to all who replied so quickly, > > > To clarify, > > I am actually wanting to remove the whole spinal column with the cord > inside, so it would be a tool to cut through the ribs attached to the > spinal column, we do a lot of pm's and are trying to make life a little > easier on the hands as cutting through the bone can be tough. > > Thanks once again > > Susan > > ---------------------------------------------------------- > Le présent message ainsi que ses éventuelles pièces jointes est > exclusivement destiné au(x) destinataire(s), personnes physiques ou > morales, qu'il désigne. > Il constitue de ce fait une correspondance à caractère privé et peut > contenir des informations confidentielles. > Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser > immédiatement l'expéditeur par retour de courrier électronique puis de le > détruire, ainsi que ses éventuelles pièces jointes, sans en conserver de > copie. > > > This message, including any attachment, is intended for the use of the > individual or entity to which it is addressed. > It is therefore to be considered as a private correspondence which may > contain confidential information. > If you are not the intended recipient, please advise the sender immediately > by reply e.mail and delete this message and any attachment thereto without > retaining a copy. > ---------------------------------------------------------- > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From gillian.2.brown <@t> gsk.com Thu Oct 20 09:35:02 2005 From: gillian.2.brown <@t> gsk.com (gillian.2.brown@gsk.com) Date: Thu Oct 20 09:34:37 2005 Subject: [Histonet] To clarify : Mechanical tool for removing spinal cord from rats. In-Reply-To: Message-ID: I regularly use a large bone and tooth saw in the UK obtainable from a materials science supplier www.materials-science-nw.co.uk If you navigate the site you'll find turbinates. I got the large one for opening up the skull of pigs and have used for opening the turbinates of Guineapigs, rats and mice. I have also used it to split the femur of a mouse LS, if this was my only use I would have gone for the smaller version though! regards Gill Brown gillian.2.brown@gsk.com Susan.Ferrigon@sanofi-aventis.com Sent by: histonet-bounces@lists.utsouthwestern.edu 20-Oct-2005 15:19 To histonet@lists.utsouthwestern.edu cc Helen.Hodgson@sanofi-aventis.com Subject [Histonet] To clarify : Mechanical tool for removing spinal cord from rats. Hi What do people use to remove spinal cords from rats during post mortems?? At present we use large scissors to cut through the bone but are looking to try a mechanical tool to replace this. Does any one have any recomendations.?? Thanks Susan Thanks to all who replied so quickly, To clarify, I am actually wanting to remove the whole spinal column with the cord inside, so it would be a tool to cut through the ribs attached to the spinal column, we do a lot of pm's and are trying to make life a little easier on the hands as cutting through the bone can be tough. Thanks once again Susan ---------------------------------------------------------- Le pr?sent message ainsi que ses ?ventuelles pi?ces jointes est exclusivement destin? au(x) destinataire(s), personnes physiques ou morales, qu'il d?signe. Il constitue de ce fait une correspondance ? caract?re priv? et peut contenir des informations confidentielles. Si ce message vous est parvenu par erreur, nous vous remercions d'en aviser imm?diatement l'exp?diteur par retour de courrier ?lectronique puis de le d?truire, ainsi que ses ?ventuelles pi?ces jointes, sans en conserver de copie. This message, including any attachment, is intended for the use of the individual or entity to which it is addressed. It is therefore to be considered as a private correspondence which may contain confidential information. If you are not the intended recipient, please advise the sender immediately by reply e.mail and delete this message and any attachment thereto without retaining a copy. ---------------------------------------------------------- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From mucram11 <@t> comcast.net Thu Oct 20 10:12:50 2005 From: mucram11 <@t> comcast.net (Pam Marcum) Date: Thu Oct 20 10:13:21 2005 Subject: [Histonet] Region II November Meeting Message-ID: <102020051512.26230.4357B3F200061AD0000066762202888744CECE030E9D0C9A03@comcast.net> Good Morning All, I wanted to remind you we have a Region II Meeting on Novermber 4th and 5th in King of Prussia PA. It is a Friday and Saturday with wide selection of speakers and topics from FNA applications to Forensic (CSI) seminars. We have preparation to classes for both QIHC (with Ethel Macrea on Friday AM) and the HT Exam (with Peggy Miccichi, Saturday all day). Please contact me for a program (we can e-mail or fax) or information about the meeting. Join us!! Pam Marcum Education Coordinator From liz <@t> premierlab.com Thu Oct 20 11:56:27 2005 From: liz <@t> premierlab.com (Elizabeth Chlipala) Date: Thu Oct 20 11:56:39 2005 Subject: [Histonet] CD11a and CD54 Message-ID: <000201c5d597$360ac2a0$a7d48a80@AMY> Hello everyone Is anyone else having problems searching the archives? I have not been able to get the search page in days. I wanted to check to see if CD11a and CD54 for mouse tissue will only work on acetone fixed frozen sections. I'm sure that this has been brought up before but unfortunately I can't get to the search page. Thanks in advance. Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC P.O. Box 18592 Boulder, Colorado 80308 Office: (303) 735-5001 Fax: (303) 735-3540 liz@premierlab.com www.premierlab.com Ship to Address: Premier Laboratory University of Colorado MCDB, Room A3B40 Boulder, Colorado 80309 From JEllin <@t> yumaregional.org Thu Oct 20 12:36:15 2005 From: JEllin <@t> yumaregional.org (Jesus Ellin) Date: Thu Oct 20 12:36:43 2005 Subject: [Histonet] Kappa, Lambda Message-ID: I have a great question out there for the IHC people. Here in the lab we are trying to work up our Kappa and lambda. The problem that most people see is the excessive background staining that comes with these two antibodies. we are treating our tissue in a H2O2 solution just before we run them on the IHC stainer (Ventanna). Here is the kicker in working them up we are using Bone marrow for control, because our pathologists would like ,control that is similar to the disease process they are looking for. Now one specimen control stains beautiful for Kappa and the other specimen control stains beautiful for Lambda. This brings me to the conclusion that the stain is working and also that different specimens no matter what stain differently. Can someone give me a universal way to make this easier on us. I know we need to block the endogenous peroxidase, and we are using H2O2.. is there anything that we can do better or is there another method for us to treat the specimens. Jesus Ellin Yuma Regional Medical Center From sharon.osborn <@t> dnax.org Thu Oct 20 12:54:45 2005 From: sharon.osborn <@t> dnax.org (Osborn, Sharon) Date: Thu Oct 20 12:55:21 2005 Subject: [Histonet] RE: histology jobs Message-ID: <29B25753F6B1D51196110002A589D444032C4859@PALMSG30.us.schp.com> Louis, I do not know what part of the country you are searching in. However, several weeks ago, I posted a job link on the histonet stating we were hiring 4 people in our general area with specific duties that could be overlapping. Visit this website link and apply for Palo Alto, CA if you would be interested in living in Northern CA San Francisco Peninsula area. http://www.schering-plough.com/schering_plough/careers/jobframe.jsp Interviews will be starting very soon. Also, check out the Advance magazine and their website for job openings. www.advanceweb.com is their link. Thirdly, there have been recruiters posting job openings on the histonet regularly; so contact some of them. Try a Google search for histology job postings. Decide on where you want to work then search directly with some of the institutions in that area. Example: I was thinking about moving to N.C. to be close to my daughter and granddaughters while my son in law was in Iraq so I looked up institutions in Raleigh, NC, the medical triangle, I found several jobs listed for Duke and other hospitals in the area. Additional thought: if you are open to travel, the temp services listed in the Advance for Medical Laboratory Professionals are often looking for people to do assignments. Often, these can lead to a permanent position. So, keep looking, set your intent and be successful...there is a shortage of good histotechs depending upon flexibility. Some places stop advertising because they have not found anyone. Success to you! sharon osborn DNAX, S-P BioPharma Palo Alto, CA Message: 4 Date: Thu, 20 Oct 2005 02:11:02 -0700 From: "Louis Apuzzio" Subject: [Histonet] Histology Opportunity To: Message-ID: Content-Type: text/plain; charset="iso-8859-1" Hello Histonetters; I am asking assistance if any one knows of any available opportunities in Histology. I am HT eligible and also a CT(ASCP). It has been difficult for me finding an available opportunity. I have excellent skills and just need an open door. Thank you all very much. Have a great day. Louis J. Apuzzio ********************************************************************* This message and any attachments are solely for the intended recipient. If you are not the intended recipient, disclosure, copying, use or distribution of the information included in this message is prohibited -- Please immediately and permanently delete. From la.sebree <@t> hosp.wisc.edu Thu Oct 20 13:59:37 2005 From: la.sebree <@t> hosp.wisc.edu (Sebree Linda A.) Date: Thu Oct 20 13:59:45 2005 Subject: [Histonet] Kappa, Lambda Message-ID: Monoclonal K and L will almost always be cleaner than polyclonal. Also, I would select several bm bxs for optimizing since some specimens are just cleaner (or dirtier) than others. Hope this helps. Linda A. Sebree University of Wisconsin Hospital & Clinics IHC/ISH Clinical & Research Laboratory DM223-VA 600 Highland Ave. Madison, WI 53792 (608)265-6596 FAX: (608)262-7174 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Jesus Ellin Sent: Thursday, October 20, 2005 12:36 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Kappa, Lambda I have a great question out there for the IHC people. Here in the lab we are trying to work up our Kappa and lambda. The problem that most people see is the excessive background staining that comes with these two antibodies. we are treating our tissue in a H2O2 solution just before we run them on the IHC stainer (Ventanna). Here is the kicker in working them up we are using Bone marrow for control, because our pathologists would like ,control that is similar to the disease process they are looking for. Now one specimen control stains beautiful for Kappa and the other specimen control stains beautiful for Lambda. This brings me to the conclusion that the stain is working and also that different specimens no matter what stain differently. Can someone give me a universal way to make this easier on us. I know we need to block the endogenous peroxidase, and we are using H2O2.. is there anything that we can do better or is there another method for us to treat the specimens. Jesus Ellin Yuma Regional Medical Center _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jkiernan <@t> uwo.ca Thu Oct 20 14:12:17 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Thu Oct 20 14:12:27 2005 Subject: [Histonet] Mechanical tool for removing spinal cord from rats. References: Message-ID: <4357EC11.DAFEC018@uwo.ca> A simple procedure is as follows: 1. Remove the head. 2. Cut through the spinal column just above the sacrum. 3. Insert the nozzle of a 2 ml syringe (filled with saline) into the spinal canal and push the plunger. 4. Catch the extruded cord (and cauda equina) in a small beaker of saline. The upper nerve roots are avulsed by this method but the histology of the cord itself is well preserved. Biochemists have used this technique for many years. It was published as "new" from an anatomical perspective 24 years ago. MEIKLE ADS & MARTIN AH 1981 A RAPID METHOD FOR REMOVAL OF THE SPINAL-CORD STAIN TECHNOLOGY 56 (4): 235-237. (According to Web of Science this paper has been cited 16 times.) Alison Meikle was an MSc student in our department, supervised by Sandy Martin. If cou need attached spinal nerves rostral to the cauda equina this is not the method to use. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ Susan.Ferrigon@sanofi-aventis.com wrote: > > Hi > > What do people use to remove spinal cords from rats during post mortems?? > At present we use large scissors to cut through the bone but are looking to > try a mechanical tool to replace this. > Does any one have any recomendations.?? > > Thanks > > Susan From mweirauch <@t> crittenton.com Thu Oct 20 14:16:47 2005 From: mweirauch <@t> crittenton.com (Maray Weirauch) Date: Thu Oct 20 14:17:15 2005 Subject: [Histonet] Formaldehyde concentration measurement Message-ID: Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray From denise.woodward <@t> uconn.edu Thu Oct 20 14:22:13 2005 From: denise.woodward <@t> uconn.edu (Woodward, Denise) Date: Thu Oct 20 14:22:23 2005 Subject: [Histonet] disposible knife holder Message-ID: Hi all, I'm looking for a used, FREE, disposable knife holder capable of fitting into the steel-knife holder of an old AO microtome (the one with the Blue/Gray outer case, not the black pre-historic model) It would be put into service in the new Histology Program at the Community College of Rhode Island. We currently have some old tempered steel knives but no knife sharpener; lots of sample disposable knives have been donated but without a holder they are useless. I figure a knife holder might be a lot easier to come by and ship than a knife sharpener! I will pay shipping for a holder, not a sharpener :-) Thanks for you help, Denise Long Woodward Educational Coordinator and Instructor of "MLCT Introduction to Histology" Community College of Rhode Island From Barry.R.Rittman <@t> uth.tmc.edu Thu Oct 20 14:41:15 2005 From: Barry.R.Rittman <@t> uth.tmc.edu (Rittman, Barry R) Date: Thu Oct 20 14:41:19 2005 Subject: [Histonet] Formaldehyde concentration measurement Message-ID: Maray Is there a specific reason that you wish to accurately measure the concentration of formaldehyde? All the formulae that I know have formalin in excess so that a difference of even several percent should make no difference. Barry -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Maray Weirauch Sent: Thursday, October 20, 2005 2:17 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Formaldehyde concentration measurement Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From DavidH <@t> marketlabinc.com Thu Oct 20 14:48:51 2005 From: DavidH <@t> marketlabinc.com (David Haagsma) Date: Thu Oct 20 14:48:59 2005 Subject: [Histonet] Formaldehyde concentration measurement Message-ID: <19E3602A16438E48B51A4250CA04B5F634C5F5@exchange.marketlab.com> Maray, Sakura has a formalin concentration test strip 5-15%. Their part number is 1463. Dave Haagsma MT(ASCP) MarketLab Inc. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Maray Weirauch Sent: Thursday, October 20, 2005 3:17 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Formaldehyde concentration measurement Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rjbuesa <@t> yahoo.com Thu Oct 20 14:49:05 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 20 14:49:14 2005 Subject: [Histonet] Kappa, Lambda In-Reply-To: Message-ID: <20051020194905.3674.qmail@web61224.mail.yahoo.com> What I will propose you probably will be an "uphill battle" with your pathologist, but the use of bone marrow bx.is a problem because they have been decalcified (presumably with a gentle EDTA protocol) and they are small and one of the things I always found very frustrating was to have a small control that will not last. I always used tonsil as control and you will have to try to convince your pathologist that if a tonsil is kappa+ and lambda+ the procedure is OK and will detect K & L+ cells anywhere. I always used monoclonal (mouse) Ig from DAKO at 1:25 dilution with HIER with citrate pH6 in a steamer. The engogenous peroxidase was quenched with 3% H2O2 X 5 minutes after HIER. Additional word of caution: controls initially + for K and L tend to weaken when the slides are kept for 4-5 weeks. According with the control slides you need, cut just enough slides for 1 week only. Rene J. "Sebree Linda A." wrote: Monoclonal K and L will almost always be cleaner than polyclonal. Also, I would select several bm bxs for optimizing since some specimens are just cleaner (or dirtier) than others. Hope this helps. Linda A. Sebree University of Wisconsin Hospital & Clinics IHC/ISH Clinical & Research Laboratory DM223-VA 600 Highland Ave. Madison, WI 53792 (608)265-6596 FAX: (608)262-7174 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Jesus Ellin Sent: Thursday, October 20, 2005 12:36 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Kappa, Lambda I have a great question out there for the IHC people. Here in the lab we are trying to work up our Kappa and lambda. The problem that most people see is the excessive background staining that comes with these two antibodies. we are treating our tissue in a H2O2 solution just before we run them on the IHC stainer (Ventanna). Here is the kicker in working them up we are using Bone marrow for control, because our pathologists would like ,control that is similar to the disease process they are looking for. Now one specimen control stains beautiful for Kappa and the other specimen control stains beautiful for Lambda. This brings me to the conclusion that the stain is working and also that different specimens no matter what stain differently. Can someone give me a universal way to make this easier on us. I know we need to block the endogenous peroxidase, and we are using H2O2.. is there anything that we can do better or is there another method for us to treat the specimens. Jesus Ellin Yuma Regional Medical Center _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From rjbuesa <@t> yahoo.com Thu Oct 20 15:00:26 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Thu Oct 20 15:00:34 2005 Subject: [Histonet] Formaldehyde concentration measurement In-Reply-To: <19E3602A16438E48B51A4250CA04B5F634C5F5@exchange.marketlab.com> Message-ID: <20051020200027.75266.qmail@web61222.mail.yahoo.com> Maray: Please remember that a formula regarded as "10% buffered formalin" solution is really a "3.7% formalin" with regards to the "pure" 37% formalin used to prepare any "10% NBF" Rene J. David Haagsma wrote: Maray, Sakura has a formalin concentration test strip 5-15%. Their part number is 1463. Dave Haagsma MT(ASCP) MarketLab Inc. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Maray Weirauch Sent: Thursday, October 20, 2005 3:17 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Formaldehyde concentration measurement Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. From travers.3 <@t> osu.edu Thu Oct 20 19:11:21 2005 From: travers.3 <@t> osu.edu (Susan Travers) Date: Thu Oct 20 16:11:56 2005 Subject: [Histonet] in situ for soluble guanyl cyclase in brain Message-ID: Has anyone had any experience (or better yet luck) in performing in situ hybridization for sGC in perfusion-fixed tissue? Any opinions would be much appreciated. ST -- Susan Travers, Ph.D. Professor, Oral Biology The Ohio State University College of Dentistry Section of Oral Biology, Room 4154 305 W. 12th Avenue Columbus, Ohio 43210-1267 (614)292-6366(V) (614)247-6945(F) From AnthonyH <@t> chw.edu.au Thu Oct 20 17:13:42 2005 From: AnthonyH <@t> chw.edu.au (Tony Henwood) Date: Thu Oct 20 17:15:54 2005 Subject: [Histonet] Formaldehyde concentration measurement Message-ID: The following might be of use: Test for validity of formalin concentrations Principle: The Schiff's reagent (PAS stain) is used to detect aldehydes and In this technique it is used to titrate a solution of formalin The concentration of which is unknown. Solutions: 1. 1 ml of formalin fixative to be tested. 2. 5 ml of 2.4% aqueous sodium bisulphite - prepare fresh. 3. Schiffs reagent. Procedure: 1. Add 1 ml of formalin to 5 ml of sodium bisulphite. 2. Mix and allow to react for 15 minutes at room temperature stirring from time to time. 3. Add 100?l of Schiff reagent. Results: If solution turns a deep violet than initial concentration of formalin is in excess of 4% (i.e. 1.6% formaldehyde). Reference: Jaspers, B., J. Histotechnol (1987) 10 (4): 263-265. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Maray Weirauch Sent: Friday, 21 October 2005 5:17 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Formaldehyde concentration measurement Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** From AnthonyH <@t> chw.edu.au Thu Oct 20 17:17:10 2005 From: AnthonyH <@t> chw.edu.au (Tony Henwood) Date: Thu Oct 20 17:17:26 2005 Subject: [Histonet] Formaldehyde concentration measurement Message-ID: Shouldn't this read as 3.7% formaldehyde not 3.7% formalin? I thought we had accepted the convention that 37%(or thereabouts) formaldehyde (concentrated formaldehyde) was 100% formalin? Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Rene J Buesa Sent: Friday, 21 October 2005 6:00 AM To: David Haagsma; Maray Weirauch; histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Formaldehyde concentration measurement Maray: Please remember that a formula regarded as "10% buffered formalin" solution is really a "3.7% formalin" with regards to the "pure" 37% formalin used to prepare any "10% NBF" Rene J. David Haagsma wrote: Maray, Sakura has a formalin concentration test strip 5-15%. Their part number is 1463. Dave Haagsma MT(ASCP) MarketLab Inc. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Maray Weirauch Sent: Thursday, October 20, 2005 3:17 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Formaldehyde concentration measurement Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** From lsunhwa <@t> gmail.com Thu Oct 20 20:16:04 2005 From: lsunhwa <@t> gmail.com (Sunhwa Lee) Date: Thu Oct 20 20:16:11 2005 Subject: [Histonet] Searching Histonet Message-ID: <177172430510201816p350aa15cr1306971090db3e73@mail.gmail.com> I would like to search for plastic embedding (Glycol Methacrylate) in Histonet. But the net seems not work. (http://www.histosearch.com/histonet.html) What's going on? And How can I use the serch tool? Sunny From dharclerode <@t> cytoritx.com Thu Oct 20 21:07:57 2005 From: dharclerode <@t> cytoritx.com (Donna Harclerode) Date: Thu Oct 20 21:07:25 2005 Subject: [Histonet] Mouse CD markers, Histosearch, apoptosis and San Diego Message-ID: <3DE0F644E093DF4BAE80C254176696A50203B7@mp-mailserver.macropore.com> Hi Liz I have been trying to search for a positive control for rat apoptosis and have not been able to get on the archives either. It is such a valuable resource it really hurts when it is not there. I never have been able to get my frozen rat hearts to work with either the Chemicon or Roche kit for TUNEL or many of the apoptosis markers (Caspase 3, PARP and DFF45). I have taken the same frozen tissue (after sectioning it for frozen), thawed it and processed into paraffin and get Fantastic TUNEL so I am going to try the other markers in paraffin. Is anyone doing apoptosis in frozen rat tissues? Is there some tick I cannot figure out? The only reason we need frozen is for CD31, which hopefully the only ab that works may available from Santa Cruz soon. I got the available lots of all their available abs. and tried them awhile back on paraffin and nothing worked. I have read on the list that they are expecting a new batch soon, but not yet. Mouse CD markers PharMingen mouse CD11a clone M17/4 works in frozen sections (1ug/ml) with polyclonal secondary. The other CD 11a clone 2D7 works at 10ug/ml, but only with the isotype specific secondary and not the polyclonal. The CD54 3E2 works well at 5ug/ml BUT you must use the hamster cocktail from PharMingen. (Jackson Immunoresearch may have come up with Syrian hamster secondaries by now, but I have not used them) I worked for PharMingen setting up the QC testing for IHC applications for their mouse, rat and human antibodies back in 1997 so I have recommended concentrations, tissues used for testing the antibodies available then and know the ones that need special secondaries. PharMingen has dropped some recommended applications (like mouse CD31, MEC13.3 for paraffin), but it worked pretty well if you used trypsin pretreatment and not microwave. The San Diego chapter of the California society has been dark for a couple years. After the Christmas holidays I am going to try and get our group up and running again. If you are in San Diego area and interested in for educational meetings, drop me and email and I will be sure to let you know what is going on. If you are willing to help get the group going again please lat me know that too. I appreciate all volunteers! Donna Harclerode, HT, (ASCP), HTL, QIHC Immunohistochemist Cytori Therapeutics 6740 Top Gun St. San Diego, CA 92121 dharclerode@cytoritx.com Is anyone else having problems searching the archives? I have not been able to get the search page in days. I wanted to check to see if CD11a and CD54 for mouse tissue will only work on acetone fixed frozen sections. I'm sure that this has been brought up before but unfortunately I can't get to the search page. Thanks in advance. Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC P.O. Box 18592 Boulder, Colorado 80308 Office: (303) 735-5001 Fax: (303) 735-3540 liz@premierlab.com www.premierlab.com aries etc. Is anyone else having problems searching the archives? I have not been able to get the search page in days. I wanted to check to see if CD11a and CD54 for mouse tissue will only work on acetone fixed frozen sections. I'm sure that this has been brought up before but unfortunately I can't get to the search page. Thanks in advance. Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC P.O. Box 18592 Boulder, Colorado 80308 Office: (303) 735-5001 Fax: (303) 735-3540 liz@premierlab.com www.premierlab.com From gu.lang <@t> gmx.at Fri Oct 21 02:34:49 2005 From: gu.lang <@t> gmx.at (Gudrun Lang) Date: Fri Oct 21 02:34:54 2005 Subject: AW: [Histonet] Formaldehyde concentration measurement In-Reply-To: Message-ID: We use 8% Formaldehyd. What would be the right amounts of reagents to test this concentration? Can you help? Gudrun Lang -----Urspr?ngliche Nachricht----- Von: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood Gesendet: Freitag, 21. Oktober 2005 00:14 An: Maray Weirauch; histonet@lists.utsouthwestern.edu Betreff: RE: [Histonet] Formaldehyde concentration measurement The following might be of use: Test for validity of formalin concentrations Principle: The Schiff's reagent (PAS stain) is used to detect aldehydes and In this technique it is used to titrate a solution of formalin The concentration of which is unknown. Solutions: 1. 1 ml of formalin fixative to be tested. 2. 5 ml of 2.4% aqueous sodium bisulphite - prepare fresh. 3. Schiffs reagent. Procedure: 1. Add 1 ml of formalin to 5 ml of sodium bisulphite. 2. Mix and allow to react for 15 minutes at room temperature stirring from time to time. 3. Add 100?l of Schiff reagent. Results: If solution turns a deep violet than initial concentration of formalin is in excess of 4% (i.e. 1.6% formaldehyde). Reference: Jaspers, B., J. Histotechnol (1987) 10 (4): 263-265. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Maray Weirauch Sent: Friday, 21 October 2005 5:17 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Formaldehyde concentration measurement Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From pmarcum <@t> vet.upenn.edu Fri Oct 21 10:42:13 2005 From: pmarcum <@t> vet.upenn.edu (Pamela Marcum) Date: Fri Oct 21 10:42:35 2005 Subject: [Histonet] Neuro and Botany Specialist Message-ID: <6.1.1.1.2.20051021112938.0198a818@mail.vet.upenn.edu> Good Morning, We have a special seminar for those who are using or will be using the Vibratome at the Region II Meeting in King of Prussia PA - November 4 and 5, 2005. It is: From Amygdala to Arabidopsis: Why's and How's of the Vibrating Microtome with Jennifer Freeland. We will have a 3 hour seminar with vibratome for demonstration and training. New and exciting ways to use the vibratome and make great sections for research. Join us by requesting a program with registration today. We are also offering a QIHC Qualification Preparation for the ASCP Registry with Ethel Macrea on Friday morning and HT (ASCP) Readiness Examination with Peggy Miccichi all day Saturday. Call or email for a copy of the program and registration for these and other great speakers and seminars. We even have a full CSI or forensic day for those interested!! Best Regards, Pamela A Marcum Manager, Histology Special Procedures University of Pennsylvania School of Veterinary Medicine R.S. Reynolds Jr. CORL New Bolton Center 382 West Street Road Kennett Square, PA 19348 Phone - 610-925-6278 Fax - 610-925-8120 E-mail - pmarcum@vet.upenn.edu or Contact: Jean Betsch, Research Specialist III Department of Otolaryngology Vestibular Research University of Pittsburgh Eye & Ear Institute, Room 146 203 Lothrop Street Pittsburgh, PA 15213 Phone: 412-647-8532 Fax: 412-647-0108 Email: jeanb@pitt.edu or betschjl@upmc.edu From rjbuesa <@t> yahoo.com Fri Oct 21 11:19:31 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Fri Oct 21 11:19:47 2005 Subject: AW: [Histonet] Formaldehyde concentration measurement Message-ID: <20051021161931.13003.qmail@web61220.mail.yahoo.com> Gudrun: Please check the posting by Tony Henwood using PAS reagent. Fisher sells Schiff reagent specific for formaldehyde detection (Cat. No.23-245687). The posted method by Tony Henwood will give you an idea but is not quantitative. If you want to quantify you will have to test different known concentrations. You could start with "pure" methanal (the "famous" 37% solution, making sure of the concentration as stated in the label. That will be your solution "A"; dilute it with deionized water in half = solution "B", and so on until you get to a dilution of ca. 0.6% after 6 consecutive dilutions. Try all of them with the method posted by Tony Henwood and prepare an absorption curve in a photocolorimeter with a "red" filter (lambda 620-720 nm) using the lambda that gives you maximum absorption. Construct an absorption curve (% absorption in the "Y axis" and "% formaldehyde" in the "X axis" (since the % in Y will depend on the % concentration in X). Any other sample could be treated with the procedure, read at the same lambda and you can find the % concentration in the graph. This value will be as inaccurate as the variation in the initial concentration. Rene J. Gudrun Lang wrote: We use 8% Formaldehyd. What would be the right amounts of reagents to test this concentration? Can you help? Gudrun Lang -----Urspr?ngliche Nachricht----- Von: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood Gesendet: Freitag, 21. Oktober 2005 00:14 An: Maray Weirauch; histonet@lists.utsouthwestern.edu Betreff: RE: [Histonet] Formaldehyde concentration measurement The following might be of use: Test for validity of formalin concentrations Principle: The Schiff's reagent (PAS stain) is used to detect aldehydes and In this technique it is used to titrate a solution of formalin The concentration of which is unknown. Solutions: 1. 1 ml of formalin fixative to be tested. 2. 5 ml of 2.4% aqueous sodium bisulphite - prepare fresh. 3. Schiffs reagent. Procedure: 1. Add 1 ml of formalin to 5 ml of sodium bisulphite. 2. Mix and allow to react for 15 minutes at room temperature stirring from time to time. 3. Add 100?l of Schiff reagent. Results: If solution turns a deep violet than initial concentration of formalin is in excess of 4% (i.e. 1.6% formaldehyde). Reference: Jaspers, B., J. Histotechnol (1987) 10 (4): 263-265. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory Manager & Senior Scientist The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145, AUSTRALIA. Tel: 612 9845 3306 Fax: 612 9845 3318 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Maray Weirauch Sent: Friday, 21 October 2005 5:17 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Formaldehyde concentration measurement Hi Histonetters, Does anyone know of a simple, inexpensive way to measure the formaldehyde concentration in a prepared, buffered formalin fixative? Thanks! Maray _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ********************************************************************** This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, the Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************** _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. From JPCOLEMA <@t> sentara.com Fri Oct 21 12:53:23 2005 From: JPCOLEMA <@t> sentara.com (JOHN COLEMAN) Date: Fri Oct 21 12:53:50 2005 Subject: [Histonet] Histo Jobs Message-ID: Louis- Quit drinking vinegar, get your HT, then call back. HT eligible= HT "ain't got". The first question on your interviews will be "are you certified?". If your answer is yes, you probably have people calling, unless you've burned some bridges. For Kappa , Lambda- I would use a multi tissue control block made in your lab by you. In this you can put a few tonsils, a few previously positive lymphomas, and both your bone marrows. It will answer all the pathoologists' questions and leave no reason to doubt your reactivity issues. I quench with 3% H2O2, use a caseinate protein block as the first two steps, and my dilutions are 1:40k and 1:15k respectively with the Dako polyclonal.(really) We retrieve 25 min in Citrate buffer 6.0 in a 98C water bath. We have clean reactions in both tonsil and bone marrow clot and bx, with some non spec staining of red cells. From JEllin <@t> yumaregional.org Fri Oct 21 13:37:14 2005 From: JEllin <@t> yumaregional.org (Jesus Ellin) Date: Fri Oct 21 13:38:01 2005 Subject: [Histonet] Kappa and Lamba Message-ID: I would like to thank everyone with there advice on this subject. It seems to me that there is a certain level of difficulty with this stain. We are seeing that the stain is working, but there is a huge amount of background on this specimen, espcially with bloody bone marrows. But there are times that the bone marrows are not bloody and we are still getting an excesive amount of staining with this procedure. But once again thank you veryone for your help and support on this issue. It is nice to know that you are not on an island alone with no rescue coming to pick you up. Jesus Ellin Yuma Regional Medical Center From ploykasek <@t> phenopath.com Fri Oct 21 13:56:26 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Fri Oct 21 13:56:55 2005 Subject: FW: [Histonet] Kappa, Lambda In-Reply-To: Message-ID: - Dear Jesus, Kappa and lambda are certainly difficult to interpret. They both can be present circulating in the serum, thus the "background" staining in most tissue sections. We use polyclonal antibodies, and get quite good results. We use a tonsil as a control as bone marrows are just too precious & small for us to use. We look for the plasma cells to be staining with a dark cytoplasmic stain & a slight "blush" to the germinal centers of the tonsil. It does take a pathologist skilled in interpreting them. A true positive is in the cytoplasm of the cells, not in-between the cells (due to circulating K/L). We do use one titer for lymph nodes, and another for bone marrows & GI biopsies. Plus a different titer & pretreatment if we're looking for amyloid. Well, I hope I've helped & not confused you. I'd be happy to do my best to answer any questions that you have. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA > I have a great question out there for the IHC people. Here in the lab we are > trying to work up our Kappa and lambda. The problem that most people see is > the excessive background staining that comes with these two antibodies. we > are treating our tissue in a H2O2 solution just before we run them on the IHC > stainer (Ventanna). Here is the kicker in working them up we are using Bone > marrow for control, because our pathologists would like ,control that is > similar to the disease process they are looking for. Now one specimen control > stains beautiful for Kappa and the other specimen control stains beautiful for > Lambda. This brings me to the conclusion that the stain is working and also > that different specimens no matter what stain differently. Can someone give > me a universal way to make this easier on us. I know we need to block the > endogenous peroxidase, and we are using H2O2.. is there anything that we can > do better or is there another method for us to treat the specimens. > > > Jesus Ellin > Yuma Regional Medical Center > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------ End of Forwarded Message This e-mail message, including any attachments, is for the sole use of the intended recipients and may contain privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message, or you may call PhenoPath Laboratories, Seattle, WA U.S.A. at (206) 374-9000. From RCHIOVETTI <@t> aol.com Fri Oct 21 14:04:59 2005 From: RCHIOVETTI <@t> aol.com (RCHIOVETTI@aol.com) Date: Fri Oct 21 14:05:18 2005 Subject: [Histonet] Searching Histonet Message-ID: <148.4ee85834.308a95db@aol.com> In a message dated 10/20/2005 6:16:49 PM US Mountain Standard Time, lsunhwa@gmail.com writes: > I would like to search for plastic embedding (Glycol Methacrylate) in > Histonet. But the net seems not work. > (http://www.histosearch.com/histonet.html) What's going on? And How > can I use the serch tool? > > Sunny, Try a little later in the day, and fill in the search box from www.histosearch.com. Usually when there's a hangup like this the server is either too busy or is having technical problems, or perhaps someone is performing maintenance on the website. I'm sure it will clear up soon. Cheers, Bob Robert (Bob) Chiovetti, Ph.D. The Microscope Works Arizona's Microscopy Resource 132 North Elster Drive Tucson, AZ 85710-3212 USA Tel./Fax 520-546-4986 Member, Arizona Small Business Association - ASBA From froyer <@t> bitstream.net Fri Oct 21 14:07:06 2005 From: froyer <@t> bitstream.net (Ford Royer) Date: Fri Oct 21 14:07:26 2005 Subject: [Histonet] Duplicate Duplicate Messages Messages In-Reply-To: <20051021175543.40CE047D82E@smtpin-1.iphouse.net> Message-ID: <007d01c5d672$a19c7b90$0b00a8c0@fords> Dear HistoNetters: Dear HistoNetters: I am receiving all postings from the HistoNet in duplicate. I am receiving all postings from the HistoNet in duplicate. Is Anyone else experiencing this phenomenon? Is Anyone else experiencing this phenomenon? I tried to unsubscribe and then re-subscribe. I tried to unsubscribe and then re-subscribe. But it did not solve the problem. But it did not solve the problem. This is beginning to annoy me. This is beginning to annoy me. I remember this happening in the past. I remember this happening in the past. But I can't remember how it was fixed. But I can't remember how it was fixed. Is there a procedure to remedy this? Is there a procedure to remedy this? Any assistance will be greatly appreciated. Any assistance will be greatly appreciated. Thanks, Thanks, ~ Ford ~ Ford Ford M. Royer, MT(ASCP) Sales Manager, Histology Product Division Minnesota Medical Specialists, Inc. 7177 Madison Ave. W. Golden Valley, MN 55427-3601 763-542-8725 phone 888-790-9686 toll free 763-546-4830 fax email web Ford M. Royer, MT(ASCP) Sales Manager, Histology Product Division Minnesota MEdical Specialists, Inc. 7177 Madison Ave. W. Golden Valley, MN 55427-3601 763-542-8725 phone 888-790-9686 toll free 763-546-4830 fax email web From kbroomal <@t> NEMOURS.ORG Fri Oct 21 14:10:57 2005 From: kbroomal <@t> NEMOURS.ORG (Kristen Broomall) Date: Fri Oct 21 14:11:19 2005 Subject: [Histonet] Duplicate Duplicate Messages Messages Message-ID: <6E41111281623B4B8A9AB8F9A7EA3437824473@wlmmsx02.nemours.org> Nope. Nope. -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Ford Royer Sent: Friday, October 21, 2005 3:07 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Duplicate Duplicate Messages Messages Dear HistoNetters: Dear HistoNetters: I am receiving all postings from the HistoNet in duplicate. I am receiving all postings from the HistoNet in duplicate. Is Anyone else experiencing this phenomenon? Is Anyone else experiencing this phenomenon? I tried to unsubscribe and then re-subscribe. I tried to unsubscribe and then re-subscribe. But it did not solve the problem. But it did not solve the problem. This is beginning to annoy me. This is beginning to annoy me. I remember this happening in the past. I remember this happening in the past. But I can't remember how it was fixed. But I can't remember how it was fixed. Is there a procedure to remedy this? Is there a procedure to remedy this? Any assistance will be greatly appreciated. Any assistance will be greatly appreciated. Thanks, Thanks, ~ Ford ~ Ford Ford M. Royer, MT(ASCP) Sales Manager, Histology Product Division Minnesota Medical Specialists, Inc. 7177 Madison Ave. W. Golden Valley, MN 55427-3601 763-542-8725 phone 888-790-9686 toll free 763-546-4830 fax email web Ford M. Royer, MT(ASCP) Sales Manager, Histology Product Division Minnesota MEdical Specialists, Inc. 7177 Madison Ave. W. Golden Valley, MN 55427-3601 763-542-8725 phone 888-790-9686 toll free 763-546-4830 fax email web _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From ploykasek <@t> phenopath.com Fri Oct 21 14:22:56 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Fri Oct 21 14:23:24 2005 Subject: [Histonet] VZV positive controls Message-ID: I'd appreciate any information on where to purchases varicella zoster positive control slides. Thanks for the help. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA This e-mail message, including any attachments, is for the sole use of the intended recipients and may contain privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message, or you may call PhenoPath Laboratories, Seattle, WA U.S.A. at (206) 374-9000. From vazquezr <@t> ohsu.edu Fri Oct 21 14:45:31 2005 From: vazquezr <@t> ohsu.edu (Robyn Vazquez) Date: Fri Oct 21 14:49:07 2005 Subject: [Histonet] Duplicate Duplicate Messages Messages Message-ID: Too bad Too bad From anh2006 <@t> med.cornell.edu Fri Oct 21 15:52:18 2005 From: anh2006 <@t> med.cornell.edu (Andrea Hooper) Date: Fri Oct 21 15:52:32 2005 Subject: [Histonet] Searching Histonet Message-ID: It hasn't been working for days for me either. From jnocito <@t> pathreflab.com Fri Oct 21 15:59:01 2005 From: jnocito <@t> pathreflab.com (Joe Nocito) Date: Fri Oct 21 15:59:18 2005 Subject: [Histonet] Block cutting protocols In-Reply-To: Message-ID: Susan, Only if you are looking for trouble. I say no Joe Nocito -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Molinari, Betsy Sent: Tuesday, October 18, 2005 5:44 AM To: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Block cutting protocols No Betsy Molinari HT (ASCP) Texas Heart Institute Cardiovascular Pathology 6770 Bertner Ave. Houston,TX 77030 832-355-6524 832-355-6812 (fax) -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Histology SLU Sent: Monday, October 17, 2005 9:05 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Block cutting protocols Hello All: I hope that you all won't "flame" me with this question. I do not want to offer any information yet as to why I am asking. I want your responses to be unbiased by the circumstances for which I am asking. The question: Do any of you techs cutting clinical paraffin embedded blocks, lay out multiple ribbons from multiple blocks on your water bath at one time and then pick up your sections for each block? I would like tech and supervisor responses please. Thanks so much. You guys are always such a help. Susan --------------------------------- Yahoo! Music Unlimited - Access over 1 million songs. Try it free. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From mhanna <@t> histosearch.com Fri Oct 21 16:17:30 2005 From: mhanna <@t> histosearch.com (Marvin Hanna) Date: Fri Oct 21 16:18:21 2005 Subject: [Histonet] Searching Histonet In-Reply-To: <148.4ee85834.308a95db@aol.com> References: <148.4ee85834.308a95db@aol.com> Message-ID: <98cfcb93548b781e38fa55a0ab4cbb56@histosearch.com> There has been a problem with the router at histosearch.com. I am working on it and the web server should be back up this evening sometime. Sorry for the inconvenience. Marvin Hanna On Oct 21, 2005, at 2:04 PM, RCHIOVETTI@aol.com wrote: > In a message dated 10/20/2005 6:16:49 PM US Mountain Standard Time, > lsunhwa@gmail.com writes: > >> I would like to search for plastic embedding (Glycol Methacrylate) in >> Histonet. But the net seems not work. >> (http://www.histosearch.com/histonet.html) What's going on? And How >> can I use the serch tool? >> >> > > Sunny, > > Try a little later in the day, and fill in the search box from > www.histosearch.com. > > Usually when there's a hangup like this the server is either too busy > or is > having technical problems, or perhaps someone is performing > maintenance on the > website. I'm sure it will clear up soon. > > Cheers, > > Bob > > Robert (Bob) Chiovetti, Ph.D. > The Microscope Works > Arizona's Microscopy Resource > 132 North Elster Drive > Tucson, AZ 85710-3212 USA > Tel./Fax 520-546-4986 > Member, Arizona Small Business Association - ASBA > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > From BSylinda <@t> aol.com Sat Oct 22 15:35:42 2005 From: BSylinda <@t> aol.com (BSylinda@aol.com) Date: Sat Oct 22 15:35:53 2005 Subject: [Histonet] Disect-Aid Message-ID: <1a6.422d78da.308bfc9e@aol.com> Hi Histoland, I am writing for more information about disect-aid. Do anyone have any experience in using this reagent for fatty tissue such as lymph node tissue. Out pathologist is interested in trying this. Any feedback of pros and cons of using this product would be greatly appreciated. Could you also forward vendor information as well. Thanks, Sylinda From rjbuesa <@t> yahoo.com Sat Oct 22 17:12:26 2005 From: rjbuesa <@t> yahoo.com (Rene J Buesa) Date: Sat Oct 22 17:12:36 2005 Subject: [Histonet] Disect-Aid In-Reply-To: <1a6.422d78da.308bfc9e@aol.com> Message-ID: <20051022221226.58568.qmail@web61214.mail.yahoo.com> Hi Sylinda: We used for lymph nodes the following solution: 100% ethanol ------2 L + Distilled water -----0.68 L + 37% formaldehyde ---0.32 L + acetic acid--0.2 L (for a total of 3.20 L). It worked very well and even the smallests lymph nodes could be detected. Rene J. BSylinda@aol.com wrote: Hi Histoland, I am writing for more information about disect-aid. Do anyone have any experience in using this reagent for fatty tissue such as lymph node tissue. Out pathologist is interested in trying this. Any feedback of pros and cons of using this product would be greatly appreciated. Could you also forward vendor information as well. Thanks, Sylinda _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. From Krat18 <@t> aol.com Sat Oct 22 21:34:41 2005 From: Krat18 <@t> aol.com (Krat18@aol.com) Date: Sat Oct 22 21:34:56 2005 Subject: [Histonet] Disect-Aid Message-ID: <146.4f5550fc.308c50c1@aol.com> Sylinda, Our PA swears by Dissect Aid ...... she uses it at both hospitals where she works. She slices the tissue into thick slices and just covers it with Dissect-Aid, and after an hour or so, the lymph nodes turn white and become very easy to spot. Dissect Aid is a product of the Decal Chemical Corporation: Please take a look at Dissect Aid on our website, www.decal-bone.com. It is a special fixative for revealing lymph nodes which we have been manufacturing for over 20 years. You can call our toll free number or reply to this email for a free sample. . Cliff Berger Decal Chemical Corp 1800-428-5856 _www.decal-bone.com_ (http://www.decal-bone.com) I hope this helps you! Karen Raterman St. Mary's Health Center St. Louis, MO 63117 _krat18@aol.com_ (mailto:krat18@aol.com) From jkiernan <@t> uwo.ca Sat Oct 22 23:01:55 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Sat Oct 22 23:02:06 2005 Subject: [Histonet] Disect-Aid References: <1a6.422d78da.308bfc9e@aol.com> Message-ID: <435B0B33.3718159A@uwo.ca> Numerous Histonet messages have extolled Davidson's fixative for this purpose. It contains less alcohol than Rene Buesa's mixture, so it's cheaper! The recipe is on Dr Bob Richmond's web site: http://members.aol.com/RSRICHMOND/histology.html For fun, and just to be pedantic: the word is dissect, and its first sy llable rhymes with hiss, not with dice. The origin is from sect (cut) and dis (apart). Until about 15 years ago it didn't matter how you pronounced the word, but there is now an important stereological technique called "the disector" (only one s) that obtains unbiased estimates of numbers and volumes of cells and other objects. The method involves collecting measurements from two (di) sections through a specimen. A dissector must obtain the specimen, and a histotechnologist (a sector who uses dyes) must prepare it for a morphometrist to apply the disector. The disector's results are tables of numbers, split up into many sections and very difficult to understand. We who dissect should proudly defend our skills by saying the word correctly. Dye-sect (for dissect) is in the same class (Homer Simpson) as nucular (for nuclear). Words like bacteria, data and media (all plural) are often used in newspapers as if they were singular rather than plural. John Kiernan Anatomy, UWO London, Canada. __________________________________________ BSylinda@aol.com wrote: > > Hi Histoland, > I am writing for more information about disect-aid. Do anyone have any > experience in using this reagent for fatty tissue such as lymph node tissue. > Out pathologist is interested in trying this. Any feedback of pros and > cons of using this product would be greatly appreciated. Could you also forward > vendor information as well. > > Thanks, > Sylinda > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From conniegrubaugh <@t> hotmail.com Sun Oct 23 14:27:52 2005 From: conniegrubaugh <@t> hotmail.com (connie grubaugh) Date: Sun Oct 23 14:28:01 2005 Subject: [Histonet] Nevada Society of Histotechnology Message-ID: The Nevada Society of Histotechnology will be having their next business and seminar Oct. 28th and 29th. 2005 There will be a Friday evening get together with the vendors. This will start at 5:30 pm. Coffee and Rolls will be provided by Fisher and Leica. Saturday morning. Saturday sign in starts at 7 am. Meeting starts at 8am. First meeting Pam Marcum-"Is Your Tissue Processor fighting you? Fight back." There will be a business meeting with lunch. Lunch will be provided by Sakura and Cardinal Health. Afternoon meetings 1 pm Sandra L. Cummings- "Putting the bug in Histotechnology. A basic understanding of Microorganisms. To Follow- Maianne Mailhoit- "Exposure control plan for Blood Borne Pathogens. This meeting is free. If you need ceu's this is the time to get them. 5.0 ceu's at this meeting. Vegas is the best place to be on Halloween weekend. Please email me with any question of call 702-396-4079 Connie Grubaugh From holling <@t> med-in.uni-saarland.de Mon Oct 24 07:15:54 2005 From: holling <@t> med-in.uni-saarland.de (holling@med-in.uni-saarland.de) Date: Mon Oct 24 07:16:08 2005 Subject: [Histonet] Staining of plastic embedded samples Message-ID: <3158.134.96.44.146.1130156154.squirrel@134.96.44.146> Hello, I?ve got a problem with staining plastic embedded samples. The samples are embedded in Technovit 9100 new. The blocks are polished and sticked on a plastic slide. With a hand saw I cut them and the slides are getting polished again and afterwards they are getting stained. Now my problem: I do the Masson Goldner or MSB stain, but the lightgreen or methyl blue doesn?t appear in the colored slice. Both are only red-brown. I don?t remove the polymer (with 2-methoxyethylacetat) before staining, because of the plastic slides. They also dissolve. I hope anybody could help me. Thanks Nicole From dford <@t> deltapathology.com Mon Oct 24 09:06:17 2005 From: dford <@t> deltapathology.com (D. Ford) Date: Mon Oct 24 09:15:48 2005 Subject: [Histonet] Current Openings for HT/HTL Message-ID: Delta Pathology, in Shreveport Louisiana, currently looking for HT/HTLs. If you have been displaced by the hurricanes and want to return to your home state or if you are looking to relocate please contact us for more information. Darlene Ford, B.S. CT, HT (ASCP) Technical Supervisor 2915 Missouri Avenue Shreveport, LA 71109 318-621-8820 phone 318-671-5922 fax dford@deltapathology.com From gcallis <@t> montana.edu Mon Oct 24 09:42:41 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Mon Oct 24 09:42:27 2005 Subject: [Histonet] Block cutting protocols In-Reply-To: <200510212103.j9LL3J8P024904@mail4.msu.montana.edu> References: <200510212103.j9LL3J8P024904@mail4.msu.montana.edu> Message-ID: <6.0.0.22.1.20051024083702.01b7bd28@gemini.msu.montana.edu> Never! Hmm, this could be a legal tissue issue, if you mix up a case or pick up the bad guys (cells as floaters) with onto a slide with next case negative good guys and the good guy gets the WRONG diagnosis, you may be seeing your attorney. Worked in a lab where this was forbidden, and we carefully skimmed the top of the waterbath between cases with a kimwipe to pick up last ribbon garbage. There is always a chance you may have a cell you don't want, but we tried to minimize the chances of this happening between all cases. At 02:59 PM 10/21/2005, you wrote: >Susan, >Only if you are looking for trouble. I say no > >Joe Nocito > >-----Original Message----- >From: histonet-bounces@lists.utsouthwestern.edu >[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Molinari, >Betsy >Sent: Tuesday, October 18, 2005 5:44 AM >To: histonet@lists.utsouthwestern.edu >Subject: RE: [Histonet] Block cutting protocols > >No > >Betsy Molinari HT (ASCP) >Texas Heart Institute >Cardiovascular Pathology >6770 Bertner Ave. >Houston,TX 77030 >832-355-6524 >832-355-6812 (fax) > > >-----Original Message----- >From: histonet-bounces@lists.utsouthwestern.edu >[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of >Histology SLU >Sent: Monday, October 17, 2005 9:05 AM >To: histonet@lists.utsouthwestern.edu >Subject: [Histonet] Block cutting protocols > >Hello All: > >I hope that you all won't "flame" me with this question. I do not want >to offer any information yet as to why I am asking. I want your >responses to be unbiased by the circumstances for which I am asking. > >The question: > >Do any of you techs cutting clinical paraffin embedded blocks, lay out >multiple ribbons from multiple blocks on your water bath at one time and >then pick up your sections for each block? > >I would like tech and supervisor responses please. Thanks so much. You >guys are always such a help. > >Susan > > >--------------------------------- > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From nsnwl <@t> neuro.hfh.edu Mon Oct 24 09:52:07 2005 From: nsnwl <@t> neuro.hfh.edu (Nancy Lemke) Date: Mon Oct 24 10:06:23 2005 Subject: [Histonet] Block cutting protocols Message-ID: <5213b638994a24b7caaeb9f7e326c0cb@neuro.hfh.edu> This is an absolute NEVER. It is similar to embedding and simultaneously opening cassettes of tissue with like specimens but different cases. Everybody knows about tissue bits flying out of the cassette when the top is removed. That is an absolute recipe for disaster. The multiple blocks from the same case on the waterbath at once is fine, of course, but different cases.......SCARY AS HALLOWEEN! Nancy Lemke Research Coordinator Hermelin Brain Tumor Center Henry Ford Hospital Detroit -----Original message----- From: "Gayle Callis" gcallis@montana.edu Date: Mon, 24 Oct 2005 10:45:25 -0400 To: "Joe Nocito" jnocito@pathreflab.com Subject: RE: [Histonet] Block cutting protocols > Never! Hmm, this could be a legal tissue issue, if you mix up a case or > pick up the bad guys (cells as floaters) with onto a slide with next case > negative good guys and the good guy gets the WRONG diagnosis, you may be > seeing your attorney. Worked in a lab where this was forbidden, and we > carefully skimmed the top of the waterbath between cases with a kimwipe to > pick up last ribbon garbage. There is always a chance you may have a cell > you don't want, but we tried to minimize the chances of this happening > between all cases. > > > > At 02:59 PM 10/21/2005, you wrote: > >Susan, > >Only if you are looking for trouble. I say no > > > >Joe Nocito > > > >-----Original Message----- > >From: histonet-bounces@lists.utsouthwestern.edu > >[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Molinari, > >Betsy > >Sent: Tuesday, October 18, 2005 5:44 AM > >To: histonet@lists.utsouthwestern.edu > >Subject: RE: [Histonet] Block cutting protocols > > > >No > > > >Betsy Molinari HT (ASCP) > >Texas Heart Institute > >Cardiovascular Pathology > >6770 Bertner Ave. > >Houston,TX 77030 > >832-355-6524 > >832-355-6812 (fax) > > > > > >-----Original Message----- > >From: histonet-bounces@lists.utsouthwestern.edu > >[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of > >Histology SLU > >Sent: Monday, October 17, 2005 9:05 AM > >To: histonet@lists.utsouthwestern.edu > >Subject: [Histonet] Block cutting protocols > > > >Hello All: > > > >I hope that you all won't "flame" me with this question. I do not want > >to offer any information yet as to why I am asking. I want your > >responses to be unbiased by the circumstances for which I am asking. > > > >The question: > > > >Do any of you techs cutting clinical paraffin embedded blocks, lay out > >multiple ribbons from multiple blocks on your water bath at one time and > >then pick up your sections for each block? > > > >I would like tech and supervisor responses please. Thanks so much. You > >guys are always such a help. > > > >Susan > > > > > >--------------------------------- > > Yahoo! Music Unlimited - Access over 1 million songs. Try it free. > >_______________________________________________ > >Histonet mailing list > >Histonet@lists.utsouthwestern.edu > >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > >_______________________________________________ > >Histonet mailing list > >Histonet@lists.utsouthwestern.edu > >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > > > > > >_______________________________________________ > >Histonet mailing list > >Histonet@lists.utsouthwestern.edu > >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > Gayle Callis > Research Histopathology Supervisor > Veterinary Molecular Biology > Montana State University - Bozeman > PO Box 173610 > Bozeman MT 59717-3610 > 406 994-6367 > 406 994-4303 (FAX) > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > ============================================================================== Go to http://henryford.com We're Henry Ford. We Can. HFHS CONFIDENTIALITY NOTICE: This email contains information from the sender that may be CONFIDENTIAL, LEGALLY PRIVILEGED, PROPRIETARY or otherwise protected from disclosure. This email is intended for use only by the person or entity to whom it is addressed. If you are not the intended recipient, any use, disclosure, copying, distribution, printing, or any action taken in reliance on the contents of this email, is strictly prohibited. If you received this email in error, please contact the sending party by replying in an email to the sender, delete the email from your computer system and shred any paper copies of the email you printed. Note to Patients: There are a number of risks you should consider before using e-mail to communicate with us. These risks are described in our Privacy Policy at http://henryford.com. Review that policy carefully before continuing to communicate with us by e-mail. 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If you do not believe that our policy gives you the privacy and security protection you need, do not send e-mail or Internet communications to us. ============================================================================== From sasa <@t> jovanovic.co.za Mon Oct 24 10:41:56 2005 From: sasa <@t> jovanovic.co.za (Sasa Jovanovic) Date: Mon Oct 24 10:42:20 2005 Subject: FW: [Histonet] Re: Immunofluorescence problem - Non-specific Message-ID: <000701c5d8b1$79582d00$640aa8c0@SinisaHome> -----Original Message----- From: Sasa Jovanovic [mailto:sasa@jovanovic.co.za] Sent: 24 October 2005 04:00 PM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] Re: Immunofluorescence problem - Non-specific Dear Ms Callis, Regarding the above mentioned subject I would like to ask if you could possible assist me with some advice......In my project I am trying to determine the presence of integrins in the rat endometrium by using immunohistochemistry on frozen cryostat sections. I am using DAKO ARK kit and was planning to add IgG2a secondary antibody as Terry Johnson suggested in the above mentioned Histonet topic.....In what stage of Dako Ark protocol and with what concentration to add this secondary antibody? Your advice will be mostly appreciated Thank you in advance Sasha From gcallis <@t> montana.edu Mon Oct 24 10:52:31 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Mon Oct 24 10:52:17 2005 Subject: [Histonet] Staining of plastic embedded samples In-Reply-To: <3158.134.96.44.146.1130156154.squirrel@134.96.44.146> References: <3158.134.96.44.146.1130156154.squirrel@134.96.44.146> Message-ID: <6.0.0.22.1.20051024090438.01b96d60@gemini.msu.montana.edu> I presume these are undecalcified bone samples, as Technovit 9100 is the methylmethacrylate version. After the final polishing of your thicker sections, etch the surface of your polished block, using an ultrasonicator, with 1% formic acid for 30 sec to 1 minute. Rinse well with running tap water, dry the section and immerse into your stains. You may have to adjust the times, but MSB may not be the ideal stain for ground, polisher MMA embedded and then acid etched sections. It may be a better stain for microtomed, thin MMA sections. You can also try etching the surface with 95 to 100 % ethanol, to soften the plastic and allow your stains to penetrate into the soft tissues but you need to do this at the beginning of staining protocol or you may remove other dyes you have applied to the section. Basically, with the acid etch, you are doing a mild surface decalcification for only a few micrometers to allow the stains to penetrate into the bone, with alcohol etch, you are only softening and NOT removing the plastic to permit these small molecular weight dyes to penetrate into the soft tissue. Whatever you do, do not over etch with acid longer than 1 min and do not use a strong mineral acid such as hydrochloric acid to etch, you will remove too much calcium at a greater depth (more micrometers) and you can have some overstained sections. If you make a mistake, you can always polish off the stain, but do not grind too much or you may remove the area you want to see. Just be aware that if you acid etch, your staining times may change with MSB. A toluidine blue stain developed for bone surface staining by Eurell and Sterchi is excellent for this purpose, as is MacNeals tetrachrome, but on the latter use an inhouse prepared stain solution, commerical stain doesn't work well at all. At 06:15 AM 10/24/2005, you wrote: >Hello, >I?ve got a problem with staining plastic embedded samples. >The samples are embedded in Technovit 9100 new. The blocks are polished >and sticked on a plastic >slide. With a hand saw I cut them and the slides are getting polished >again and afterwards they >are getting stained. >Now my problem: I do the Masson Goldner or MSB stain, but the lightgreen >or methyl blue doesn?t >appear in the colored slice. Both are only red-brown. >I don?t remove the polymer (with 2-methoxyethylacetat) before staining, >because of the plastic >slides. They also dissolve. > >I hope anybody could help me. > >Thanks Nicole > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From doninn <@t> mail.nih.gov Mon Oct 24 10:50:56 2005 From: doninn <@t> mail.nih.gov (Donin, Nick (NIH/NCI)) Date: Mon Oct 24 10:59:22 2005 Subject: [Histonet] Frozen Section Protocol Message-ID: <16A0583FB1644E4DB8C0A0265028B6FD02B4E815@nihexchange13.nih.gov> Histonet, My PI has requested that I section and stain sections of a subcutaneous human tumor that is frozen. He wants me to cut 10 micron sections, and then stain the tumor and look at it using fluorescence (as opposed to some chromogen technique like Vectastain). Can anyone suggest a protocol? Below is the protocol I have been using so far, with poor results (high background, very little specific staining. Thanks everyone. Freeze tumor in isopentane - 5 seconds Section tumor at 10 microns and place on slide Wash 1x with PBS Fix 15 min with 4% PFA Block 1 hr with 5% NGS with 0.3% Triton Incubate 1 hr RT with primary antibody Wash 3x 5 min with PBS Incubate 1 hr RT with secondary antibody Wash 3x 5 min with PBS Apply Dapi and coverslip (all staining and washing is done by covering tissue with solution while tissue is on the glass slide) Nick Donin CRTA Neuro-Oncology Branch National Cancer Institute National Institutes of Health 9000 Rockville Pike Building 35, Room 2B-203 Bethesda, MD 20892 From cfavara <@t> niaid.nih.gov Mon Oct 24 11:26:08 2005 From: cfavara <@t> niaid.nih.gov (Favara, Cynthia (NIH/NIAID)) Date: Mon Oct 24 11:26:18 2005 Subject: [Histonet] Frozen Section Protocol Message-ID: Your question leads me to believe you are using an antibody that you have used previously with good results and a direct HRP tagged secondary. If this is indeed the case you should be able to adjust the concentrations so that both secondaries have the same concentration. It would be necessary to include a no primary control to determine the amount of fluorescence due to the tagged secondary. I like to do a control for auto fluorescence as well. I did not hear any reference to a known positive or negative control those would have an impact on determining the protocol as well. I personally would cut a bit thinner 6-8um. c Cynthia Favara NIAID/NIH/RML/LPVD 903 South 4th Street Hamilton, MT 59840 406-363-9317 Disclaimer: The information in this e-mail and any of its attachments is confidential and may contain sensitive information. It should not be used by anyone who is not the original intended recipient. If you have received this e-mail in error please inform the sender and delete it from your mailbox or any other storage devices. National Institute of Allergy and Infectious Diseases shall not accept liability for any statements made that are sender's own and not expressly made on behalf of the NIAID by one of its representatives -----Original Message----- From: Donin, Nick (NIH/NCI) Sent: Monday, October 24, 2005 8:51 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Frozen Section Protocol Histonet, My PI has requested that I section and stain sections of a subcutaneous human tumor that is frozen. He wants me to cut 10 micron sections, and then stain the tumor and look at it using fluorescence (as opposed to some chromogen technique like Vectastain). Can anyone suggest a protocol? Below is the protocol I have been using so far, with poor results (high background, very little specific staining. Thanks everyone. Freeze tumor in isopentane - 5 seconds Section tumor at 10 microns and place on slide Wash 1x with PBS Fix 15 min with 4% PFA Block 1 hr with 5% NGS with 0.3% Triton Incubate 1 hr RT with primary antibody Wash 3x 5 min with PBS Incubate 1 hr RT with secondary antibody Wash 3x 5 min with PBS Apply Dapi and coverslip (all staining and washing is done by covering tissue with solution while tissue is on the glass slide) Nick Donin CRTA Neuro-Oncology Branch National Cancer Institute National Institutes of Health 9000 Rockville Pike Building 35, Room 2B-203 Bethesda, MD 20892 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From PMonfils <@t> Lifespan.org Mon Oct 24 12:25:06 2005 From: PMonfils <@t> Lifespan.org (Monfils, Paul) Date: Mon Oct 24 12:25:18 2005 Subject: [Histonet] GFP in Paraffin Sections? Message-ID: <09C945920A6B654199F7A58A1D7D1FDE017175FB@lsexch.lsmaster.lifespan.org> Does anyone know if GFP (Green Fluorescent Protein) maintains its fluorescence through paraffin processing? Can the fluorescence be viewed in an otherwise unstained paraffin section? After deparaffinization, should the section be coverslipped with an aqueous medium, or the standard medium used for paraffin sections? From Dorothy.L.Webb <@t> HealthPartners.Com Mon Oct 24 12:54:05 2005 From: Dorothy.L.Webb <@t> HealthPartners.Com (Webb, Dorothy L) Date: Mon Oct 24 12:54:15 2005 Subject: [Histonet] Alk 1 Message-ID: <0E394B648E5284478A6CCB78E5AFDA2718C47F@hpes1.HealthPartners.int> We use the Alk-1 antibody from cell marque on the Benchmark and have very good results. Any other info needed, contact me direct Email. ________________________________________ This e-mail and any files transmitted with it are confidential and are intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient or the individual responsible for delivering the e-mail to the intended recipient, please be advised that you have received this e-mail in error and that any use, dissemination, forwarding, printing, or copying of this e-mail is strictly prohibited. If you have received this e-mail in error, please immediately notify the HealthPartners Support Center by telephone at (952) 967-6600. You will be reimbursed for reasonable costs incurred in notifying us. From vazquezr <@t> ohsu.edu Mon Oct 24 13:00:36 2005 From: vazquezr <@t> ohsu.edu (Robyn Vazquez) Date: Mon Oct 24 13:00:59 2005 Subject: [Histonet] Block cutting protocols Message-ID: I still wouldn't even from the same case. Just opening the door... Robyn OHSU From mclarke <@t> allsaintshealthcare.org Mon Oct 24 13:06:33 2005 From: mclarke <@t> allsaintshealthcare.org (Clarke, Mary) Date: Mon Oct 24 13:06:48 2005 Subject: [Histonet] DISSECT AID Message-ID: <6FE2A453DA437F4D96FAAF363F20ABB20FC8B3@WFEXBE04.wfsi.priv> WE are using the Dissect Aid and tow of my pathologists really like it. Terri Clarke Histology Supervisor All Saints Laboratory 3801 Spring Street Racine, Wi 53405 mclarke@allsaintshealthcare.org 262-687-6609 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of histonet-request@lists.utsouthwestern.edu Sent: Sunday, October 23, 2005 12:01 PM To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 23, Issue 27 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit http://lists.utsouthwestern.edu/mailman/listinfo/histonet or, via email, send a message with subject or body 'help' to histonet-request@lists.utsouthwestern.edu You can reach the person managing the list at histonet-owner@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than "Re: Contents of Histonet digest..." Today's Topics: 1. Disect-Aid (BSylinda@aol.com) 2. Re: Disect-Aid (Rene J Buesa) 3. Re: Disect-Aid (Krat18@aol.com) 4. Re: Disect-Aid (John Kiernan) ---------------------------------------------------------------------- Message: 1 Date: Sat, 22 Oct 2005 16:35:42 EDT From: BSylinda@aol.com Subject: [Histonet] Disect-Aid To: histonet@lists.utsouthwestern.edu Message-ID: <1a6.422d78da.308bfc9e@aol.com> Content-Type: text/plain; charset="US-ASCII" Hi Histoland, I am writing for more information about disect-aid. Do anyone have any experience in using this reagent for fatty tissue such as lymph node tissue. Out pathologist is interested in trying this. Any feedback of pros and cons of using this product would be greatly appreciated. Could you also forward vendor information as well. Thanks, Sylinda ------------------------------ Message: 2 Date: Sat, 22 Oct 2005 15:12:26 -0700 (PDT) From: Rene J Buesa Subject: Re: [Histonet] Disect-Aid To: BSylinda@aol.com, histonet@lists.utsouthwestern.edu Message-ID: <20051022221226.58568.qmail@web61214.mail.yahoo.com> Content-Type: text/plain; charset=iso-8859-1 Hi Sylinda: We used for lymph nodes the following solution: 100% ethanol ------2 L + Distilled water -----0.68 L + 37% formaldehyde ---0.32 L + acetic acid--0.2 L (for a total of 3.20 L). It worked very well and even the smallests lymph nodes could be detected. Rene J. BSylinda@aol.com wrote: Hi Histoland, I am writing for more information about disect-aid. Do anyone have any experience in using this reagent for fatty tissue such as lymph node tissue. Out pathologist is interested in trying this. Any feedback of pros and cons of using this product would be greatly appreciated. Could you also forward vendor information as well. Thanks, Sylinda _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. ------------------------------ Message: 3 Date: Sat, 22 Oct 2005 22:34:41 EDT From: Krat18@aol.com Subject: Re: [Histonet] Disect-Aid To: BSylinda@aol.com, histonet@lists.utsouthwestern.edu Message-ID: <146.4f5550fc.308c50c1@aol.com> Content-Type: text/plain; charset="US-ASCII" Sylinda, Our PA swears by Dissect Aid ...... she uses it at both hospitals where she works. She slices the tissue into thick slices and just covers it with Dissect-Aid, and after an hour or so, the lymph nodes turn white and become very easy to spot. Dissect Aid is a product of the Decal Chemical Corporation: Please take a look at Dissect Aid on our website, www.decal-bone.com. It is a special fixative for revealing lymph nodes which we have been manufacturing for over 20 years. You can call our toll free number or reply to this email for a free sample. . Cliff Berger Decal Chemical Corp 1800-428-5856 _www.decal-bone.com_ (http://www.decal-bone.com) I hope this helps you! Karen Raterman St. Mary's Health Center St. Louis, MO 63117 _krat18@aol.com_ (mailto:krat18@aol.com) ------------------------------ Message: 4 Date: Sun, 23 Oct 2005 00:01:55 -0400 From: John Kiernan Subject: Re: [Histonet] Disect-Aid To: BSylinda@aol.com Cc: histonet@lists.utsouthwestern.edu Message-ID: <435B0B33.3718159A@uwo.ca> Content-Type: text/plain; charset=us-ascii Numerous Histonet messages have extolled Davidson's fixative for this purpose. It contains less alcohol than Rene Buesa's mixture, so it's cheaper! The recipe is on Dr Bob Richmond's web site: http://members.aol.com/RSRICHMOND/histology.html For fun, and just to be pedantic: the word is dissect, and its first sy llable rhymes with hiss, not with dice. The origin is from sect (cut) and dis (apart). Until about 15 years ago it didn't matter how you pronounced the word, but there is now an important stereological technique called "the disector" (only one s) that obtains unbiased estimates of numbers and volumes of cells and other objects. The method involves collecting measurements from two (di) sections through a specimen. A dissector must obtain the specimen, and a histotechnologist (a sector who uses dyes) must prepare it for a morphometrist to apply the disector. The disector's results are tables of numbers, split up into many sections and very difficult to understand. We who dissect should proudly defend our skills by saying the word correctly. Dye-sect (for dissect) is in the same class (Homer Simpson) as nucular (for nuclear). Words like bacteria, data and media (all plural) are often used in newspapers as if they were singular rather than plural. John Kiernan Anatomy, UWO London, Canada. __________________________________________ BSylinda@aol.com wrote: > > Hi Histoland, > I am writing for more information about disect-aid. Do anyone have any > experience in using this reagent for fatty tissue such as lymph node tissue. > Out pathologist is interested in trying this. Any feedback of pros and > cons of using this product would be greatly appreciated. Could you also forward > vendor information as well. > > Thanks, > Sylinda > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------ _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet End of Histonet Digest, Vol 23, Issue 27 **************************************** Privileged/Confidential information may be contained in this message. The information contained in this message is intended only for the use of the recipient(s) named above and their co-workers who are working on the same matter. The recipient of this information is prohibited from disclosing the information to any other party unless this disclosure has been authorized in advance. 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From sandosis <@t> uia.net Mon Oct 24 13:23:35 2005 From: sandosis <@t> uia.net (sandosis@uia.net) Date: Mon Oct 24 13:23:45 2005 Subject: [Histonet] Desmin that will stain neoplastic muscle? Message-ID: <200510241823.j9OINau81341@smtp1.uia.net> Can someone recommend a source for Desmin that will stain the neoplastic muscle cells? Currently we are using the pre-dilute from Ventana. (DE-R-11) This only seems to stain mature, normal muscle. Thanks much, Sandra C. Orange, CA ` --------------------------------------------- This message was sent using the UIA Web Mail Server. ULTIMATE Internet Access, Inc http://www.uia.net/ From Rcartun <@t> harthosp.org Mon Oct 24 14:09:21 2005 From: Rcartun <@t> harthosp.org (Richard Cartun) Date: Mon Oct 24 14:09:50 2005 Subject: [Histonet] Desmin that will stain neoplastic muscle? Message-ID: I use DakoCytomation's monoclonal (clone D33). Richard Richard W. Cartun, Ph.D. Director, Immunopathology & Histology Assistant Director, Anatomic Pathology Hartford Hospital 80 Seymour Street Hartford, CT 06102 (860) 545-1596 (860) 545-0174 Fax >>> 10/24/05 02:23PM >>> Can someone recommend a source for Desmin that will stain the neoplastic muscle cells? Currently we are using the pre-dilute from Ventana. (DE-R-11) This only seems to stain mature, normal muscle. Thanks much, Sandra C. Orange, CA ` --------------------------------------------- This message was sent using the UIA Web Mail Server. ULTIMATE Internet Access, Inc http://www.uia.net/ _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ----------------------------------------------- Confidentiality Notice This e-mail message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential or proprietary information which is legally privileged. Any unauthorized review, use, disclosure, or distribution is prohibited. If you are not the intended recipient, please promptly contact the sender by reply e-mail and destroy all copies of the original message. From pmarcum <@t> vet.upenn.edu Mon Oct 24 14:39:07 2005 From: pmarcum <@t> vet.upenn.edu (Pamela Marcum) Date: Mon Oct 24 14:39:21 2005 Subject: [Histonet] Region II Meeting November 4 & 5 in King of Prussia PA Message-ID: <6.1.1.1.2.20051024152653.019d0d78@mail.vet.upenn.edu> Good Afternoon All, Ever wonder what a CSI technician actually does and how? Come to our meeting and learn from members of the New Jersey State Highway Patrol Crime Lab. They will explain what is real and what is just good writing. Find out how high profile cases are handled and why the stress is so high for both the Medical Examiner who gets the body and then the CSI laboratory with the evidence. You can also learn about mummies too!!! We have management seminars for those who are new in supervising a laboratory or those who feel you can never learn enough about how to manage your people. Contact us for a program and registration information. We still have some room and would love more company for the meeting!! If you love to shop or need to for the coming season the hotel has a shuttle to the King of Prussia Plaza and Court Shopping Center. (One of the largest in the US) Please contact: Jean Betsch, Research Specialist III Department of Otolaryngology Vestibular Research University of Pittsburgh Eye & Ear Institute, Room 146 203 Lothrop Street Pittsburgh, PA 15213 Phone: 412-647-8532 Fax: 412-647-0108 Email: jeanb@pitt.edu or betschjl@upmc.edu Best Regards, Pamela A Marcum Manager, Histology Special Procedures University of Pennsylvania School of Veterinary Medicine R.S. Reynolds Jr. CORL New Bolton Center 382 West Street Road Kennett Square, PA 19348 Phone - 610-925-6278 Fax - 610-925-8120 E-mail - pmarcum@vet.upenn.edu From Barb.Richmond <@t> mckennan.org Mon Oct 24 14:52:31 2005 From: Barb.Richmond <@t> mckennan.org (Barb Richmond) Date: Mon Oct 24 14:52:39 2005 Subject: [Histonet] "dirty slides" Message-ID: I'm wondering if anyone has a secret on how to keep our H&E slides free of paraffin and fingerprints. Our procedure is to check the label and section quality with the block after the slide has been coverslipped and labeled. We end up with paraffin on the slides, so we have to wipe them down with alcohol. Any other suggestions?? From brett_connolly <@t> merck.com Mon Oct 24 16:44:40 2005 From: brett_connolly <@t> merck.com (Connolly, Brett M) Date: Mon Oct 24 16:45:19 2005 Subject: [Histonet] PCNA versus Ki67 Message-ID: <355C35514FEAC9458F75947F5270974D079684@usctmx1103.merck.com> I have been a big proponent of using Ki67 instead of PCNA as a more accurate proliferation marker after reading that PCNA may in fact carry over into G0. Recently, some of my colleagues have been clamoring for PCNA. I'd be interested to hear which marker other labs are using. Thanks, Brett Brett M. Connolly, Ph.D. Merck & Co., Inc. MRL, Imaging Research WP-44K PO Box 4 West Point, PA 19486 PH 215-652-2501 fax. 215-993-6803 e-mail. brett_connolly@merck.com ------------------------------------------------------------------------------ Notice: This e-mail message, together with any attachments, contains information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates (which may be known outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD and in Japan, as Banyu) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system. ------------------------------------------------------------------------------ From gcallis <@t> montana.edu Mon Oct 24 16:52:18 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Mon Oct 24 16:52:34 2005 Subject: [Histonet] GFP in Paraffin Sections? In-Reply-To: <09C945920A6B654199F7A58A1D7D1FDE017175FB@lsexch.lsmaster.l ifespan.org> References: <09C945920A6B654199F7A58A1D7D1FDE017175FB@lsexch.lsmaster.lifespan.org> Message-ID: <6.0.0.22.1.20051024154740.01b40be8@gemini.msu.montana.edu> Paul, You may have success or you may not. We cannot maintain our eGFP with paraffin processing, or with any frozen section fixed with solvents. Alcohols may ruin the GFP and its fluorescent properties, however there are those who have had success. One can always come back with antiGFP (Rockland makes are very nice goat antiGFP) and then detect the goat antiGFP with a Donkey antiGoat-FITC. It is certainly worth a try to do what you suggest and try both ways or mounting medias. If your tissues are FFPE, then you will be battling green autofluorescence too. I have a review of autofluorescence with GFP in mind I am sending to you via private email, a very nice publication. At 11:25 AM 10/24/2005, you wrote: >Does anyone know if GFP (Green Fluorescent Protein) maintains its >fluorescence through paraffin processing? Can the fluorescence be viewed in >an otherwise unstained paraffin section? After deparaffinization, should >the section be coverslipped with an aqueous medium, or the standard medium >used for paraffin sections? > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From cfavara <@t> niaid.nih.gov Mon Oct 24 17:03:53 2005 From: cfavara <@t> niaid.nih.gov (Favara, Cynthia (NIH/NIAID)) Date: Mon Oct 24 17:04:02 2005 Subject: [Histonet] GFP in Paraffin Sections? Message-ID: Good advice from Gayle. I have yet to be successful her with GFAP on frozens and need to come back with an anti -GFP. The most sensitive antibody I have found is from Roche, cocktail of 2 monoclonal mouse Ab's. I have not had a problem with mouse on mouse as we are usually dealing with neuronal tissue. Jamie Erickson did a poster around 98 at NSH and will send you her conclusions. You could look in the archives for her information. c Cynthia Favara NIAID/NIH/RML/LPVD 903 South 4th Street Hamilton, MT 59840 406-363-9317 Disclaimer: The information in this e-mail and any of its attachments is confidential and may contain sensitive information. It should not be used by anyone who is not the original intended recipient. If you have received this e-mail in error please inform the sender and delete it from your mailbox or any other storage devices. National Institute of Allergy and Infectious Diseases shall not accept liability for any statements made that are sender's own and not expressly made on behalf of the NIAID by one of its representatives -----Original Message----- From: Monfils, Paul [mailto:PMonfils@Lifespan.org] Sent: Monday, October 24, 2005 10:25 AM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] GFP in Paraffin Sections? Does anyone know if GFP (Green Fluorescent Protein) maintains its fluorescence through paraffin processing? Can the fluorescence be viewed in an otherwise unstained paraffin section? After deparaffinization, should the section be coverslipped with an aqueous medium, or the standard medium used for paraffin sections? _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Adesupod <@t> aol.com Mon Oct 24 18:24:03 2005 From: Adesupod <@t> aol.com (Adesupod@aol.com) Date: Mon Oct 24 18:24:13 2005 Subject: [Histonet] B-5 FIXATIVE Message-ID: <6d.501e4e9d.308ec713@aol.com> Hi, Pls I will like to know how many of you guys are still fixing your Bone Marrow biopsies in B-5 Fixative. Is it even safe to fix tissue specimens in B-5 fixative? Hoping to read from you asap. Adesupo. From emry <@t> u.washington.edu Mon Oct 24 19:03:42 2005 From: emry <@t> u.washington.edu (Trisha Emry) Date: Mon Oct 24 19:02:44 2005 Subject: [Histonet] Bob Coma? Message-ID: Looking for Bob Coma of Leica. email address? tel number? Thanks, Trisha From MVaughan4 <@t> ucok.edu Mon Oct 24 20:03:57 2005 From: MVaughan4 <@t> ucok.edu (MVaughan4@ucok.edu) Date: Mon Oct 24 20:04:28 2005 Subject: [Histonet] How do you get red pulp really red? In-Reply-To: Message-ID: Hello everyone, I teach a histology class and often have the students make their own slides. I notice that my very old histology slides have very bright red pulp and dark white pulp. I assume they were prepared using H&E staining. I have tried but the red is never as bright as the old slides; the hematoxylin part is fine however. I use eosin in 70% EtOH. Any suggestions as to how I can get the red pulp red? Thanks, I like perusing the topics. Mel Melville B. Vaughan, Ph. D. Assistant Professor Department of Biology University of Central Oklahoma Edmond, OK 73034 From Krat18 <@t> aol.com Mon Oct 24 22:58:56 2005 From: Krat18 <@t> aol.com (Krat18@aol.com) Date: Mon Oct 24 22:59:12 2005 Subject: [Histonet] How do you get red pulp really red? Message-ID: <24.7bb9aff1.308f0780@aol.com> I'd suggest using Eosin Y with Phloxine, from Sigma Chemical Company, Fisher Scientific or Richard-Allan Scientific, among others. That's what our pathologists like as a counterstain with hematoxylin because it gives a brighter red than the Eosin Y alone. I hope this is what you need. Karen Raterman _krat18@aol.com_ (mailto:krat18@aol.com) From Krat18 <@t> aol.com Mon Oct 24 23:04:07 2005 From: Krat18 <@t> aol.com (Krat18@aol.com) Date: Mon Oct 24 23:04:19 2005 Subject: [Histonet] B-5 FIXATIVE Message-ID: <5b.74a34aa1.308f08b7@aol.com> We have switched to B-Plus Fix from BBC Biochemical or MarketLab. Our docs are happy with it, and it contains no mercury. Karen Raterman _krat18@aol.com_ (mailto:krat18@aol.com) From vanann702 <@t> skmc.gov.ae Mon Oct 24 23:13:20 2005 From: vanann702 <@t> skmc.gov.ae (Anne Van Binsbergen) Date: Mon Oct 24 23:11:55 2005 Subject: [Histonet] How do you get red pulp really red? Message-ID: <0C44F1AAEE47D54DA4210A60AB206F5E02F6D680@SKMCEMAIL.skmc.gov.ae> Hi Mel Eosin in 70% alc.... - what % eosin - is it eosin Y or G I use a working solution of roughly 0,5% eosin Y in dist H2O to which I add a few drops of acetic acid and a 'dash' of 1% aq phloxine For frozens we use a 1% with a few drops of acetic acid - faster Its really all a matter of timing, which also depends on staining method (manual, auto, etc) - trial and error should get you there Remember that it isn't always the one stain/colour (Haem or Eos) which is the culprit - it's the balance between them If your water/section is too alkaline the eosin will not stain the section - so wash really well after 'blueing' When I took over this lab they were using alc eosin and the Paths hated the staining Now they LOVE it. Annieinarabia -----Original Message----- From: MVaughan4@ucok.edu [mailto:MVaughan4@ucok.edu] Sent: Tuesday, October 25, 2005 5:04 AM To: Subject: [Histonet] How do you get red pulp really red? Hello everyone, I teach a histology class and often have the students make their own slides. I notice that my very old histology slides have very bright red pulp and dark white pulp. I assume they were prepared using H&E staining. I have tried but the red is never as bright as the old slides; the hematoxylin part is fine however. I use eosin in 70% EtOH. Any suggestions as to how I can get the red pulp red? Thanks, I like perusing the topics. Mel Melville B. Vaughan, Ph. D. Assistant Professor Department of Biology University of Central Oklahoma Edmond, OK 73034 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From ncoolen <@t> burns.nl Tue Oct 25 05:40:32 2005 From: ncoolen <@t> burns.nl (Coolen, Neeltje) Date: Tue Oct 25 05:40:42 2005 Subject: [Histonet] brdU incorporation in keratinocytes Message-ID: <8A67A7CA4D84D44CB1B8F3D657CE599E3004BF@server-02.brandwonden.local> Dear Histonetters, I am trying to detect brdU incorporation in keratinocytes, but the keratinocytes which are not pulsed with brdU also stained positive for brdU. This was the protocol I used: The cells were pulsed with medium containing 100uM brdU (Sigma, cat. no. B5002) or with medium alone for 3h, fixed in 4% formaldehyde at 4oC for 30 min, incubated in HCl (1N) for 10 min on ice followed by incubation in HCl (2N) for 10 min at r.t before moving them to an incubator for 20 min at 37oC. Immediately after the acid, cells were washed in 0.1 M sodium borate (pH 8.5) for 10 min, incubated with 5% normal rabbit serum for 30 min and incubated with anti-brdU antibody (1:50, MP Biomedicals, clone IIB5) for 1h at r.t. As a secondary antibody I used biotinylated rabbit anti-mouse Ig (DAKO) and I used streptavidine alexa fluor 488 (Molecular Probes). Does anyone have a solution for this problem? Thank you for your help. Neeltje Coolen -------------------------------------- N.A. Coolen, PhD student VSBN Zeestraat 26 1941 AJ Beverwijk The Netherlands tel: +31 251 275 564 From ree3 <@t> leicester.ac.uk Tue Oct 25 06:09:27 2005 From: ree3 <@t> leicester.ac.uk (Edwards, R.E.) Date: Tue Oct 25 06:09:36 2005 Subject: [Histonet] E: beta-2- microglobulin Message-ID: Looking for an antibody to the above that works, as ever, on paraffin processed mouse tissues... Many thanks Richard Edwards MRC TOX UNIT LEICESTER...U.K.... ___ From jerry.santiago <@t> jax.ufl.edu Tue Oct 25 06:35:45 2005 From: jerry.santiago <@t> jax.ufl.edu (Santiago, Jerry) Date: Tue Oct 25 06:39:27 2005 Subject: [Histonet] Florida Call for Abstract Message-ID: <6B072F080D8FE048BCB9F51780D1FB631C89C2@jaxmail.umc.ufl.edu> The Florida Society for Histotechnology is now accepting abstracts for their meeting to be held on May 5-7, 2006 in Deerfield Beach, Florida. All abstracts should be sent to: Jerry Santiago 658 Southland Lane Orange Park, FL 32065 or e-mailed to: jerry.santiago@jax.ufl.edu From settembr <@t> umdnj.edu Tue Oct 25 08:43:01 2005 From: settembr <@t> umdnj.edu (Dana Settembre) Date: Tue Oct 25 08:46:34 2005 Subject: [Histonet] PCNA versus Ki67 Message-ID: We use Ki67 and have rarely used PCNA. What is "go"? Is that a phase? Dana Settembre University Hospital - UDMNJ, Newark, NJ >>> "Connolly, Brett M" 10/24/2005 5:44:40 PM >>> I have been a big proponent of using Ki67 instead of PCNA as a more accurate proliferation marker after reading that PCNA may in fact carry over into G0. Recently, some of my colleagues have been clamoring for PCNA. I'd be interested to hear which marker other labs are using. Thanks, Brett Brett M. Connolly, Ph.D. Merck & Co., Inc. MRL, Imaging Research WP-44K PO Box 4 West Point, PA 19486 PH 215-652-2501 fax. 215-993-6803 e-mail. brett_connolly@merck.com ------------------------------------------------------------------------------ Notice: This e-mail message, together with any attachments, contains information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates (which may be known outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD and in Japan, as Banyu) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system. ------------------------------------------------------------------------------ _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From mgr <@t> seattleskincancer.com Tue Oct 25 09:12:05 2005 From: mgr <@t> seattleskincancer.com (Manager) Date: Tue Oct 25 09:18:36 2005 Subject: [Histonet] Bob Coma? In-Reply-To: Message-ID: <20051025141815.7EBBC6DC57@smtp2.pacifier.net> He is with Bartels & Stout. 425-453-1705. Beth -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Trisha Emry Sent: Monday, October 24, 2005 5:04 PM To: histo Subject: [Histonet] Bob Coma? Looking for Bob Coma of Leica. email address? tel number? Thanks, Trisha _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From anne.lewin <@t> bms.com Tue Oct 25 09:16:48 2005 From: anne.lewin <@t> bms.com (Anne C Lewin) Date: Tue Oct 25 09:23:58 2005 Subject: [Histonet] PCNA versus Ki67 In-Reply-To: References: Message-ID: <435E3E50.4010703@bms.com> We also routinely use Ki67, although have gotten some good qualitative results with PCNA. The different phases of the cell cycle are: G1 (growth and chromosome preparation), S (formation of centrosomes, synthesis of DNA - prophase, metaphase, anaphase and telophase), G2 (Preparation for mitosis), and M (mitosis). G0(zero, not O), is when the cell "steps out" of the cell cycle and isn't actively proliferating. This is why it would be misleading for purposes of analysis for the IHC to include these cells. -Anne Lewin Dana Settembre wrote: >We use Ki67 and have rarely used PCNA. What is "go"? Is that a >phase? >Dana Settembre >University Hospital - UDMNJ, Newark, NJ > > > >>>>"Connolly, Brett M" 10/24/2005 5:44:40 >>>> >>>> >PM >>> >I have been a big proponent of using Ki67 instead of PCNA as a more >accurate >proliferation marker after reading that PCNA may in fact carry over >into G0. >Recently, some of my colleagues have been clamoring for PCNA. > >I'd be interested to hear which marker other labs are using. > >Thanks, >Brett > >Brett M. Connolly, Ph.D. >Merck & Co., Inc. >MRL, Imaging Research >WP-44K >PO Box 4 >West Point, PA 19486 >PH 215-652-2501 >fax. 215-993-6803 >e-mail. brett_connolly@merck.com > > > >------------------------------------------------------------------------------ >Notice: This e-mail message, together with any attachments, contains >information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, >New Jersey, USA 08889), and/or its affiliates (which may be known >outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD >and in Japan, as Banyu) that may be confidential, proprietary >copyrighted and/or legally privileged. It is intended solely for the use >of the individual or entity named on this message. If you are not the >intended recipient, and have received this message in error, please >notify us immediately by reply e-mail and then delete it from your >system. >------------------------------------------------------------------------------ > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > From brett_connolly <@t> merck.com Tue Oct 25 09:36:49 2005 From: brett_connolly <@t> merck.com (Connolly, Brett M) Date: Tue Oct 25 09:37:37 2005 Subject: [Histonet] PCNA versus Ki67 Message-ID: <355C35514FEAC9458F75947F5270974D079689@usctmx1103.merck.com> I guess my font made it look like 'go' , it's really G 'zero' as Anne Lewin explained. Brett -----Original Message----- From: Dana Settembre [mailto:settembr@umdnj.edu] Sent: Tuesday, October 25, 2005 9:43 AM To: histonet@lists.utsouthwestern.edu; Connolly, Brett M Subject: Re: [Histonet] PCNA versus Ki67 We use Ki67 and have rarely used PCNA. What is "go"? Is that a phase? Dana Settembre University Hospital - UDMNJ, Newark, NJ >>> "Connolly, Brett M" 10/24/2005 5:44:40 PM >>> I have been a big proponent of using Ki67 instead of PCNA as a more accurate proliferation marker after reading that PCNA may in fact carry over into G0. Recently, some of my colleagues have been clamoring for PCNA. I'd be interested to hear which marker other labs are using. Thanks, Brett Brett M. Connolly, Ph.D. Merck & Co., Inc. MRL, Imaging Research WP-44K PO Box 4 West Point, PA 19486 PH 215-652-2501 fax. 215-993-6803 e-mail. brett_connolly@merck.com ---------------------------------------------------------------------------- -- Notice: This e-mail message, together with any attachments, contains information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates (which may be known outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD and in Japan, as Banyu) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system. ---------------------------------------------------------------------------- -- _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------------------------------------------------------ Notice: This e-mail message, together with any attachments, contains information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates (which may be known outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD and in Japan, as Banyu) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system. ------------------------------------------------------------------------------ From vazquezr <@t> ohsu.edu Tue Oct 25 10:10:05 2005 From: vazquezr <@t> ohsu.edu (Robyn Vazquez) Date: Tue Oct 25 10:10:33 2005 Subject: [Histonet] Bob Coma? Message-ID: I know a Bob Coma of Bartles and Stout? 1-425-453-1705 in Bellevue, WA Robyn OHSU From gcallis <@t> montana.edu Tue Oct 25 11:37:23 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Tue Oct 25 11:37:40 2005 Subject: [Histonet] New product to solve autofluorescence woes Message-ID: <6.0.0.22.1.20051025102745.01b6fae8@gemini.msu.montana.edu> Dear H'netters If you are doing immunofluorescence staining on fixed and permeabilized cells and tissues and suffer from autofluorescence, there are new things on the horizon. A new product is out although I have not tried it, but am curious to know if it works. Molecular Probes now has Image-IT signal enhancer, cat#136933 which is applied before you start your staining. It would certainly be worth a try for those who use paraformaldehyde or NBF fixation, and possibly for FFPE tissue sections destined for immunofluorescence staining. and maybe for PFA fixed frozen sections with GFP? Contact their tech services if you have any questions about this new product and you can read about it online at www.probes.invitrogen.com ------ I think it would be worth trying. If anyone has used it , please post your results - it would be a huge help to others performing immunofluorescent staining and suffering from autofluorescence woes to know if this works. Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From NSEARCY <@t> swmail.sw.org Tue Oct 25 11:43:59 2005 From: NSEARCY <@t> swmail.sw.org (Nita Searcy) Date: Tue Oct 25 11:44:01 2005 Subject: [Histonet] Reagent Rental Agreements Message-ID: Have just heard that in above, one can't be "awarded" instrument - that its against Medicare rules. I have had acquisitions like it in the past, what has changed ? I was told that "fair market value" has to be paid at the end of contract. Any thoughts? Have you experienced such a problem? How do vendors / institutions get around this problem? Thanks From MadaryJ <@t> MedImmune.com Tue Oct 25 12:18:07 2005 From: MadaryJ <@t> MedImmune.com (Madary, Joseph) Date: Tue Oct 25 12:22:06 2005 Subject: [Histonet] locking paraffin block cabinets Message-ID: <746FDB897740814EA52BDDCB5ED1DDBC027D8B05@medimmune4.medimmune.com> My pathological wants to get some locking paraffin block cabinets. Is there such an animal out there? In my last lab we had a GLP archive and had the cabinets of blocks inside a locking cabinet. Here we do not have the space but still need the security. Nick Madary, HT/HTL(ASCP)QIHC Histology, Medimmune Inc Gaithersburg, MD 20878 301.398.6113 From Mara.Fairman <@t> RoperSaintFrancis.com Tue Oct 25 12:24:50 2005 From: Mara.Fairman <@t> RoperSaintFrancis.com (Fairman Mara) Date: Tue Oct 25 12:25:03 2005 Subject: [Histonet] re- B-5 FIXATIVE Message-ID: We have used B Plus fixative (from BBC) for a number of years and our pathologists like it. We use it for bone marrows and lymph nodes. Our facility is going "mercury free". Mara Fairman, HT(ASCP) BHS Anatomic Pathology Supervisor Roper Hospital 316 Calhoun Street Charleston, S.C. 29401 843-724-2264 Fax- 843-724-2356 Beeper 764-1489 mara.fairman@ropersaintfrancis.com Our Mission: Healing all people with compassion, faith and excellence Confidentiality Notice: This e-mail message, including any attachments, is for the sole use of the intended recipients and may contain confidential and/or legally privileged information. If you are not the intended recipient, please contact the sender by reply e-mail and destroy all copies of the original message. Any unauthorized review, use, disclosure, or distribution is prohibited. Thank you. From jqb7 <@t> cdc.gov Tue Oct 25 13:01:03 2005 From: jqb7 <@t> cdc.gov (Bartlett, Jeanine) Date: Tue Oct 25 13:04:45 2005 Subject: [Histonet] locking paraffin block cabinets Message-ID: I know of two: Sakura's Lab Aid Ultra and Lab Storage System's High Density Cabinet Jeanine Bartlett, HT(ASCP) Centers for Disease Control and Prevention 1600 Clifton Road, MS/G-32 Atlanta, GA 30333 (404) 639-3590 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Madary, Joseph Sent: Tuesday, October 25, 2005 1:18 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] locking paraffin block cabinets My pathological wants to get some locking paraffin block cabinets. Is there such an animal out there? In my last lab we had a GLP archive and had the cabinets of blocks inside a locking cabinet. Here we do not have the space but still need the security. Nick Madary, HT/HTL(ASCP)QIHC Histology, Medimmune Inc Gaithersburg, MD 20878 301.398.6113 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From TJJ <@t> Stowers-Institute.org Tue Oct 25 13:46:43 2005 From: TJJ <@t> Stowers-Institute.org (Johnson, Teri) Date: Tue Oct 25 13:46:52 2005 Subject: [Histonet] Re: BrdU incorporation in keratinocytes Message-ID: Not having done BrdU in cultured cells, I may be way off. I would suspect your antibody might need to be titered out further and/or you may want to try detecting the antibody using an anti-mouse Alexa 488 instead of a biotin-streptavidin system. Also, that's quite a hearty denaturing protocol you are using. Do you have a publication that recommends doing it that way? I don't know if that is related to the false positive staining, but it's certainly worth looking at. Teri Johnson Managing Director Histology Facility Stowers Institute for Medical Research 1000 E. 50th St. Kansas City, MO 64133 From histology <@t> gradymem.org Tue Oct 25 15:03:45 2005 From: histology <@t> gradymem.org (histology@gradymem.org) Date: Tue Oct 25 15:04:11 2005 Subject: [Histonet] CAP question on grossing Message-ID: CAP question ANP.11610: If specimens are dissected by individuals other than a pathologist or pathology resident, do such individuals qualify as high complexity testing personnel under CLIA-88 regulations? My question: Does the dissection of any kind of tissue count as high complexity testing? --even gallbladders, appendix, skin, etc? How about specimens that do not require any dissection but are submitted in whole--such as small biopsies, TURP's, etc?...Do individuals have to be CLIA-88 qualified to put biopsies in? If an individual does meet the CLIA-88 qualifications, does he have to have a certificate from CLIA? Thanks for any response. Our CAP inspection is coming up in a couple of months. Angie Barnett, HTL(ASCP) Grady Memorial Hospital Pathology Department 405/224-2258 histology@gradymem.org From Charles.Embrey <@t> carle.com Tue Oct 25 16:26:26 2005 From: Charles.Embrey <@t> carle.com (Charles.Embrey) Date: Tue Oct 25 16:24:41 2005 Subject: [Histonet] CAP question on grossing Message-ID: Yes, any grossing of tissue is high complexity testing. No, CLIA does not issue certificates. Charles Embrey PA(ASCP) -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of histology@gradymem.org Sent: Tuesday, October 25, 2005 3:04 PM To: Histonet Subject: [Histonet] CAP question on grossing CAP question ANP.11610: If specimens are dissected by individuals other than a pathologist or pathology resident, do such individuals qualify as high complexity testing personnel under CLIA-88 regulations? My question: Does the dissection of any kind of tissue count as high complexity testing? --even gallbladders, appendix, skin, etc? How about specimens that do not require any dissection but are submitted in whole--such as small biopsies, TURP's, etc?...Do individuals have to be CLIA-88 qualified to put biopsies in? If an individual does meet the CLIA-88 qualifications, does he have to have a certificate from CLIA? Thanks for any response. Our CAP inspection is coming up in a couple of months. Angie Barnett, HTL(ASCP) Grady Memorial Hospital Pathology Department 405/224-2258 histology@gradymem.org _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From tyler-wellington <@t> northwestern.edu Tue Oct 25 16:38:49 2005 From: tyler-wellington <@t> northwestern.edu (Tyler W.) Date: Tue Oct 25 16:38:56 2005 Subject: [Histonet] newfound chatter Message-ID: <20051025213849.7EE429E844@merle.it.northwestern.edu> While the chatter on this list is always delightful, the chatter that I have seen recently in my sections is less than desirable. Sorry, once again, for the seemingly rookie question, but I was wandering why, all of a sudden, this has become a problem. What adjustments would one make to the blade angle? Does hydrating the block have a lot to do with it? I cut down the hydration time recently because overhydrating caused my sections not to ribbon. I also switched to surgipath formula r paraffin. Does anyone have experience with this type (or, by all means others) and good ways to eliminate chatter? Just curious, thank you. Bonus question: What's your favorite type of paraffin? Thanks again--Tyler W. From cbass <@t> bidmc.harvard.edu Tue Oct 25 16:46:46 2005 From: cbass <@t> bidmc.harvard.edu (Caroline Bass) Date: Tue Oct 25 16:47:00 2005 Subject: [Histonet] does fixation kill native EGFP or RFP fluorescence? Message-ID: Hello, My boss insists that EGFP or RFP expressing tissue will lose the native fluorescence if fixed. Is this true? I know it isn't true for tissue culture, but he is very insistent that it will occur. I have cut the liver into blocks and immersion fixed them in 10%NBF overnight at 4 degrees celcius.. If I cut a thin section of fresh tissue I can see a lot of signal. He insists I am wasting my time by sectioning the blocks. Any advice would be appreciated. Thanks, Caroline From liz <@t> premierlab.com Tue Oct 25 16:54:32 2005 From: liz <@t> premierlab.com (Elizabeth Chlipala) Date: Tue Oct 25 16:55:01 2005 Subject: [Histonet] deOlmos Amino Cupric Siver Stain Message-ID: <000001c5d9ae$b1fea640$a7d48a80@AMY> Is anyone familiar with this stain and if so do you happen to have a protocol that you are willing to share. Thanks in Advance Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC P.O. Box 18592 Boulder, Colorado 80308 Office: (303) 735-5001 Fax: (303) 735-3540 liz@premierlab.com www.premierlab.com Ship to Address: Premier Laboratory University of Colorado MCDB, Room A3B40 Boulder, Colorado 80309 From c.m.vanderloos <@t> amc.uva.nl Wed Oct 26 03:01:41 2005 From: c.m.vanderloos <@t> amc.uva.nl (C.M. van der Loos) Date: Wed Oct 26 03:01:53 2005 Subject: [Histonet] RE: PCNA versus Ki67 Message-ID: Dear Brett, As been said before, you will find more positive cells with PCNA than with Ki67. As far as I know is due to a different half-life of these antigens. That's the reason why also cells in the G0 phase might be PCNA positive. But, if you're in doubt whether to use PCNA or Ki67, why not visualizing both of them in double staining? Since you have a mouse anti-PCNA and rabbit anti-Ki67 (LabVision/Neomarkers) life is simple here. Prepare a cocktail of these primaries (appropriately diluted of course). Than, prepare a 1:1 cocktail of PowerVision-anti-rabbit/AP and PowerVision-anti-mouse/HRP (ImmunoVision) and incubate for 60 min. Develop AP and HRP activity respectively in blue (Vector Blue or home-made Fast Blue BB) and red (AEC or Vector NovaRed). Done!!! Cheers, Chris van der Loos, PhD Dept. of Pathology Academic Medical Center M2-230 Meibergdreef 9 NL-1105 AZ Amsterdam The Netherlands phone: +31 20 5665631 fax: +31 20 6960389 Date: Mon, 24 Oct 2005 17:44:40 -0400 From: "Connolly, Brett M" Subject: [Histonet] PCNA versus Ki67 To: histonet@lists.utsouthwestern.edu I have been a big proponent of using Ki67 instead of PCNA as a more accurate proliferation marker after reading that PCNA may in fact carry over into G0. Recently, some of my colleagues have been clamoring for PCNA. I'd be interested to hear which marker other labs are using. Thanks, Brett Brett M. Connolly, Ph.D. Merck & Co., Inc. MRL, Imaging Research WP-44K PO Box 4 West Point, PA 19486 PH 215-652-2501 fax. 215-993-6803 From LJApuzzio <@t> msn.com Wed Oct 26 06:11:53 2005 From: LJApuzzio <@t> msn.com (Louis Apuzzio) Date: Wed Oct 26 06:12:10 2005 Subject: [Histonet] An Offer of Gratitude Message-ID: Hello Histonetters; I would like to thank each and everyone, for their suggestions and aid for seeking an opportunity in Histology. There has been positive responses, in addition to some negativity. My search still continues and I thought all was well, within a 24 hour period. I had been offered a position, in the private sector, resume' sent, letters of recommendation were also used and lastly, active managers were available to discuss my character and performance. All was too good, then the bottom dropped out. I was packed and ready to move forward, then the big concern of not being "certified". I am not in search of a " free meal ticket'" and desire only to satisfy my goal of becoming HT registered. One has to be gainfully employed, within the field, to obtain the necessary materials to satisfy the BOR. I only ask that an opportunity, with an open door be granted to satisfy my personal quest and also to be a team member. LJApuzzio@msn.com From johanna.jackson <@t> csc.mrc.ac.uk Wed Oct 26 06:57:37 2005 From: johanna.jackson <@t> csc.mrc.ac.uk (Jackson, Johanna) Date: Wed Oct 26 06:58:09 2005 Subject: [Histonet] Eosin-Hematoxylin solution according to Ehrlich(from Sigma) Message-ID: <2A1F3E27491CBF46BC14ACB3E5C7D89A0BD8B6@icex3.ic.ac.uk> Hi, I need to stain frozen rat brain sections (15um) with H&E. As I'm new to this I bought the ready-made Eosin-Hematoxylin solution according to Ehrlich from Fluka/Sigma. Does anyone have a protocol for using ready-made H&E solutions? It seems a bit confusing as I can't do the steps, such as washing etc, that usually go between the haematoxylin and eosin. Many thanks, Jo Stem Cell Imaging MRC Clinical Sciences Centre Imperial College London Hammersmith Hospital Campus Du Cane Road London W12 0NN From gu.lang <@t> gmx.at Wed Oct 26 07:23:10 2005 From: gu.lang <@t> gmx.at (Gudrun Lang) Date: Wed Oct 26 07:23:15 2005 Subject: [Histonet] questions about rhodanine-copper stain Message-ID: I am dealing with the rhodanine stain, because we shall establish it in our histolab. I've read three different protocols (AFIP, Churukian, webpath). AFIP and Churukian require sodiumacetat solutions to mix with the stocksolution, the others require just the dilution of the stocksolution in Aqua dest. Why? Do both versions work? Churukian demands coverslipping with aqueous medium, the others don't. Is the staining result soluble in ethanol? The stability of the 0,2% rhodanine/ethanol stocksolution is described between one day and three months. What is the right shelf live? I hope someone can explain it to me. Gudrun Lang General hospital Linz, Austria From nellisk <@t> mail.nih.gov Wed Oct 26 08:24:21 2005 From: nellisk <@t> mail.nih.gov (Nellis, Kevin Lee (NIH/NCI)) Date: Wed Oct 26 08:24:37 2005 Subject: [Histonet] HISTOPATHOLOGY TECHNICIAN RECRUITMENT at NCI Message-ID: <27C204BD76CBC142BA1AE46D62A8548E18D9E9C9@nihexchange9.nih.gov> Dear all, Please circulate this job announcement information. The National Cancer Institute is recruiting a HISTOPATHOLOGY TECHNICIAN, GS-0646-09/10 (SALARY RANGE: 43,365.00 - 62,086.00 USD per year). RELOCATION EXPENSES AND RECRUITMENT BONUS MAY BE PAID. Please see Job Announcement Number NCI-05-99225 on the USAJOBS.GOV website for full details. http://jobsearch.usajobs.opm.gov/getjob.asp?JobID=35321937 Applications will be accepted from US Citizens, from current and former competitive service Federal employees, and people eligible under special hiring authorities, through November 7, 2005. Please read the job posting carefully and submit your resume and answer KSAs for full consideration. Cordially, Kevin L. Nellis, MS, MT(ASCP) Clinical Laboratory Manager Laboratory of Pathology, NCI From vazquezr <@t> ohsu.edu Wed Oct 26 09:01:08 2005 From: vazquezr <@t> ohsu.edu (Robyn Vazquez) Date: Wed Oct 26 09:01:44 2005 Subject: [Histonet] HISTOPATHOLOGY TECHNICIAN RECRUITMENT at NCI Message-ID: And where is this job located? From nellisk <@t> mail.nih.gov Wed Oct 26 09:07:23 2005 From: nellisk <@t> mail.nih.gov (Nellis, Kevin Lee (NIH/NCI)) Date: Wed Oct 26 09:07:38 2005 Subject: [Histonet] HISTOPATHOLOGY TECHNICIAN RECRUITMENT at NCI Message-ID: <27C204BD76CBC142BA1AE46D62A8548E18D9E9CC@nihexchange9.nih.gov> Robyn: The job is located in Bethesda Maryland, on the campus of the National Institutes of Health. Hope this answers your question. All the details are in the job announcement. Thanks, Kevin From Charles.Embrey <@t> carle.com Wed Oct 26 09:48:51 2005 From: Charles.Embrey <@t> carle.com (Charles.Embrey) Date: Wed Oct 26 09:49:04 2005 Subject: [Histonet] CAP question on grossing Message-ID: CAP and CLIA have never made a separation in "transference" from grossing. Ever CLIA statement I have ever read on the subject states that grossing any tissue is high complexity testing. "Transference" is not even recognized in the CAP checklist. I routinely assist on CAP inspections and I would love to see a lab try to sell me on "transference". Of course it is only a Phase II write up but if the problem is not documented as corrected the lab can lose its accreditation. Liability and appropriate pay are other considerations I personally have mentioned in other e-mails but have been flamed for. My personal feeling is that if someone wants to undervalue their talent and work that is up to them. As long as labs can get techs to do part of the pathologists job for about a tenth of the pathologist's salary, those people will be prayed upon. Charles Embrey, PA(ASCP) -----Original Message----- From: Jesus Ellin [mailto:JEllin@yumaregional.org] Sent: Tuesday, October 25, 2005 7:36 PM To: Charles.Embrey Subject: RE: [Histonet] CAP question on grossing What about transference, our pathologist's are saying that small bx's or anything that does not require cutting or manipulation falls under this category. They are saying any registered or registry eligable can do this portion of the gross. What needs to happen is have stated black in white what techs can or can not do. This not only puts a lot of liability on the tech, it also is having tech do a PA or pathologist job at a techs wage, which is not right at all. Were can someone find out what the scale is for grossing tech's? I know it depends on the institution, but there has to be some sort of support for this other than requirements that CLIA and CAP set forth. Also what other than a phase two what can also happen if a facility is caught doing this.?? Jesus Ellin Yuma Regional Medical Center From jkiernan <@t> uwo.ca Wed Oct 26 09:56:26 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Wed Oct 26 09:56:37 2005 Subject: [Histonet] Eosin-Hematoxylin solution according to Ehrlich(from Sigma) References: <2A1F3E27491CBF46BC14ACB3E5C7D89A0BD8B6@icex3.ic.ac.uk> Message-ID: <435F991A.FB6095E9@uwo.ca> Are you sure you have the right name for the solution? Ehrlich's hematoxylin is an air-oxidized hemalum with a large excess of alum. The original 1886 formulation is Water 100 ml Absolute alcohol 100 ml Glycerol 100 ml Glacial acetic acid 10 ml Hematoxylin 2g Aluminum potassium sulfate (alum) "to excess" (presumably saturation). Leave it for a month before using; it keeps for years. This is the original recipe, published as a half-page note in Z. wiss. Mikrosk. 3:150 (1886). It's more easily available in "The Collected Papers of Paul Ehrlich", ed. F. Himmelweit, 1956, Vol. 1 p.113. London: Pergamon Press. I've made and used it and can vouch for its efficacy as a selective progressive nuclear stain. The recipes for Ehrlich's hematoxylin given in books commonly have less alum than the original, and are consequently somewhat more rapidly acting. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ Eosin Y is applied as a counterstain following a hemalum nuclear stain. I don't think you could mix the two together because eosin Y and related dyes are almost insoluble at the low pH of a hemalum solution. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ "Jackson, Johanna" wrote: > > Hi, > > I need to stain frozen rat brain sections (15um) with H&E. As I'm new to this I bought the ready-made Eosin-Hematoxylin solution according to Ehrlich from Fluka/Sigma. Does anyone have a protocol for using ready-made H&E solutions? It seems a bit confusing as I can't do the steps, such as washing etc, that usually go between the haematoxylin and eosin. > > Many thanks, > > Jo > > > Stem Cell Imaging > MRC Clinical Sciences Centre > Imperial College London > Hammersmith Hospital Campus > Du Cane Road > London > W12 0NN > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From gcallis <@t> montana.edu Wed Oct 26 10:25:49 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Wed Oct 26 10:26:04 2005 Subject: [Histonet] does fixation kill native EGFP or RFP fluorescence? In-Reply-To: References: Message-ID: <6.0.0.22.1.20051026085851.01b57f88@gemini.msu.montana.edu> Caroline, Are you going frozen sections or are you going on with paraffin processing? By saying you are cutting a fresh section, I presumeyou are snap freezing a tissue without any fixation prior to the freezing part. OR are you snap freezing the NBF fixed liver? and do you cryoprotect with 25% - 30% sucrose after fixation then snap freeze? By a lot of signal, do you mean everywhere or specific for cells? You could be seeing autofluorescence - one of the problems with viewing eGFP after NBF or PFA fixation. NBF or paraformaldehyde will preserve these proteins, but we always snap froze fresh tissue, then fixed the sections in PFA, 2% for only a few minutes. We had much better signal if we simply cut a frozen section, let it air dry, rinse very gently with DPBS, and coverslip the UNFIXED section with Molecular Probes Prolong Gold antifade, ready to use (hard set) in order to protect the fluorescence. There is no law to say you have to fix a frozen section to see eGFP, and we do this with DsRED also. The joy is you have no aldehyde induced autofluorescence, merely the natural autofluorescing components found in liver. We do look at these unfixed sections on day of cryotomy, it takes a bit of coordination but it is another way to see eGFP. Beware of lipofuscins (hey, did I say that correctly?) This was discussed recently on Histonet and you can go to Archives with keyword search to read up on this. We cannot maintain eGFP with any organic solvent exposure i.e. frozen section fixed with acetone or acetone alcohol mixture. When we used paraformaldehyde, we had best signal with shortest fixation time and low concentration of PFA, i.e. 2 min fixation of a frozen section versus a 10 min fixation of a FS in cold 2% PFA in DPBS but our autofluorescence was a pain. One way to prove you are seeing eGFP is do an antiGFP on an adjacent section, Rockland Goat antiGFP is superb, and you can do this with immunofluorescence or IHC. I suggest you go to the Clontech website and get the Living Colours Manual in PDF form, it has a wealth of information for eGFP, etc. on fixation, problems encountered and what will autofluoresce in tissues. Good luck At 03:46 PM 10/25/2005, you wrote: >Hello, > >My boss insists that EGFP or RFP expressing tissue will lose the >native fluorescence if fixed. Is this true? I know it isn't true >for tissue culture, but he is very insistent that it will occur. > >I have cut the liver into blocks and immersion fixed them in 10%NBF >overnight at 4 degrees celcius.. If I cut a thin section of fresh >tissue I can see a lot of signal. He insists I am wasting my time by >sectioning the blocks. > >Any advice would be appreciated. > >Thanks, > >Caroline > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From nsnwl <@t> neuro.hfh.edu Wed Oct 26 10:41:58 2005 From: nsnwl <@t> neuro.hfh.edu (Nancy Lemke) Date: Wed Oct 26 10:42:13 2005 Subject: [Histonet] whole brain sections Message-ID: Histonetters, Does anyone have an idea where I could find instructions for using an A/O-Reichert Tetrander (sp?) microtome for whole brain sections? The only citation I can easily find is from 1919 and in German. I know there must be something somewhere that can give me some in depth information and probably there are many of you that have first hand experience with such a beast. Thanks in advance, Nancy Lemke Research Coordinator Hermelin Brian Tumor Center Henry Ford Hospital Detroit, MI ============================================================================== Go to http://henryford.com We're Henry Ford. We Can. HFHS CONFIDENTIALITY NOTICE: This email contains information from the sender that may be CONFIDENTIAL, LEGALLY PRIVILEGED, PROPRIETARY or otherwise protected from disclosure. This email is intended for use only by the person or entity to whom it is addressed. If you are not the intended recipient, any use, disclosure, copying, distribution, printing, or any action taken in reliance on the contents of this email, is strictly prohibited. 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If you do not believe that our policy gives you the privacy and security protection you need, do not send e-mail or Internet communications to us. ============================================================================== From JMahoney <@t> alegent.org Wed Oct 26 11:20:15 2005 From: JMahoney <@t> alegent.org (Janice A Mahoney) Date: Wed Oct 26 11:21:17 2005 Subject: [Histonet] CAP question on grossing Message-ID: I agree with Charles. I have given labs CAP deficiencies for this when I have been on inspection teams. If you count it, measure it, describe it or cut it you are grossing according to the feedback I have received from the CLIA folks (CAP follows CLIA's lead on this). Read the CLIA rule. If personnel have been performing grossing prior to a specified date and you have documentation of their annual competency and training you may be ok. If not, you are putting your lab at risk. Jan Mahoney Omaha, NE >>> "Charles.Embrey" 10/26/2005 9:48 AM >>> CAP and CLIA have never made a separation in "transference" from grossing. Ever CLIA statement I have ever read on the subject states that grossing any tissue is high complexity testing. "Transference" is not even recognized in the CAP checklist. I routinely assist on CAP inspections and I would love to see a lab try to sell me on "transference". Of course it is only a Phase II write up but if the problem is not documented as corrected the lab can lose its accreditation. Liability and appropriate pay are other considerations I personally have mentioned in other e-mails but have been flamed for. My personal feeling is that if someone wants to undervalue their talent and work that is up to them. As long as labs can get techs to do part of the pathologists job for about a tenth of the pathologist's salary, those people will be prayed upon. Charles Embrey, PA(ASCP) -----Original Message----- From: Jesus Ellin [mailto:JEllin@yumaregional.org] Sent: Tuesday, October 25, 2005 7:36 PM To: Charles.Embrey Subject: RE: [Histonet] CAP question on grossing What about transference, our pathologist's are saying that small bx's or anything that does not require cutting or manipulation falls under this category. They are saying any registered or registry eligable can do this portion of the gross. What needs to happen is have stated black in white what techs can or can not do. This not only puts a lot of liability on the tech, it also is having tech do a PA or pathologist job at a techs wage, which is not right at all. Were can someone find out what the scale is for grossing tech's? I know it depends on the institution, but there has to be some sort of support for this other than requirements that CLIA and CAP set forth. Also what other than a phase two what can also happen if a facility is caught doing this.?? Jesus Ellin Yuma Regional Medical Center _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From STapper <@t> slhduluth.com Wed Oct 26 12:03:45 2005 From: STapper <@t> slhduluth.com (Tapper, Sheila) Date: Wed Oct 26 12:03:56 2005 Subject: [Histonet] TBS Shurmark Plus Message-ID: <9A3C36FB3D39404398867DFE2A4B5DE7010967A5@slhw2smail01.slhdomain.com> I am looking for someone that is using the slide unit from TBS the newest model the Shurmark Plus. I have seen a demo at NSH, but am looking for real users. Thanks! Sheila Tapper HT(ASCP) St. Luke's Hospital Duluth, MN This communication is intended for the use of the person or entity to whom it is addressed and may contain information that is privileged and confidential, the disclosure of which is governed by applicable law. If the reader of this message is not the intended recipient, or the employee or agent responsible for delivering it to the intended recipient, you are hereby notified that any dissemination, distribution, or copying of this information is prohibited. If you have received this message in error, please notify sender immediately. From JosefaNava <@t> texashealth.org Wed Oct 26 12:10:57 2005 From: JosefaNava <@t> texashealth.org (Nava, Josefa) Date: Wed Oct 26 12:11:07 2005 Subject: [Histonet] OCT 3/4 Antibody for Seminoma Message-ID: <2C515C1049EAF5459EFD8C9B929078A4194551@phdex03.txhealth.org> Hello everyone, Where can I order the Antibody OCT3/4 that will work on our Ventana Machines? I appreciate any information . Josie Nava The information contained in this message and any attachments is intended only for the use of the individual or entity to which it is addressed, and may contain information that is PRIVILEGED, CONFIDENTIAL, and exempt from disclosure under applicable law. If you are not the intended recipient, you are prohibited from copying, distributing, or using the information. Please contact the sender immediately by return e-mail and delete the original message from your system. From anh2006 <@t> med.cornell.edu Wed Oct 26 12:27:05 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Wed Oct 26 12:22:17 2005 Subject: [Histonet] Colon mets - cytokeratin IHC Message-ID: Hi All, I am looking for the best possible IHC stain to use on colon met samples to distinguish single met cells amongst normal adjacent liver. I was thinking AE1/AE3 or some CK20 antibody or any others the experts may suggest. I would love to get suggestions of best clones and sources for the antibodies as well as staining dilutions and protocols if possible. Thanks, Andrea -- From anh2006 <@t> med.cornell.edu Wed Oct 26 12:28:19 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Wed Oct 26 12:23:30 2005 Subject: [Histonet] Red HRP chromagen/substrate Message-ID: I am looking for the best red chromagen/substrate for HRP out there. Suggestions and sources/catalog numbers needed! Thanks, Andrea -- From weneng2004 <@t> yahoo.com Wed Oct 26 12:36:10 2005 From: weneng2004 <@t> yahoo.com (wen eng) Date: Wed Oct 26 12:36:19 2005 Subject: [Histonet] CTGF IHC Message-ID: <20051026173610.25016.qmail@web53406.mail.yahoo.com> Hello histonetters, I was asked to do CTGF IHC on mouse paraffin sections. Does anybody have idea which antibody is good? It will be highly appreciated if you can share your procedure. Thanks in advance, Wen --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. From gcallis <@t> montana.edu Wed Oct 26 12:46:10 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Wed Oct 26 12:46:22 2005 Subject: [Histonet] Re: Red HRP chromagen/substrate In-Reply-To: References: Message-ID: <6.0.0.22.1.20051026113739.01b8d220@gemini.msu.montana.edu> Andrea, Try DAKO's AEC+, ready to use.#K3461 for 15 ml or K3469 for 110 ml. However Dr. Chris van der Loos's in house recipe is excellent and much less pricey, just as sensitive as AEC+. If you want the recipe, I will be happy to send what Chris shared with me. It also depends on the color red you want, AEC is a reddish orange, while VECTOR NovaRed, is equivalent to DAB has a deeper brick red color. We prefer the truer red of AEC to the the deeper brick red. At 11:28 AM 10/26/2005, you wrote: >I am looking for the best red chromagen/substrate for HRP out there. >Suggestions and sources/catalog numbers needed! > >Thanks, >Andrea >-- > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From gcallis <@t> montana.edu Wed Oct 26 12:53:13 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Wed Oct 26 12:53:25 2005 Subject: [Histonet] Cholesterol staining suggestions Message-ID: <6.0.0.22.1.20051026114957.01b4d488@gemini.msu.montana.edu> I had an inquiry about what staining method is best for cholesterol but after looking at the methods (PAN) in the literature I have on hand, I am not sure I want to deal with this type of staining protocol. Would it be better to do an immunostain? If so, this would be on murine tissue. Any suggestions are welcome for the grad student asking about cholesterol Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From Karen.Heckford <@t> CHW.edu Wed Oct 26 12:57:15 2005 From: Karen.Heckford <@t> CHW.edu (Heckford, Karen - SMMC-SF) Date: Wed Oct 26 12:57:23 2005 Subject: [Histonet] Expired Stains Message-ID: Hi Everyone, I was wondering if anyone knows how JCAHO views expired Special Stains. I have some expired Special Stains. We do low volume, trying to use up some stains before they expire can be difficult. I can still get the stains to work and the Pathologists like them. Should I throw them away and order new? As you know the stains can be costly. Thanks for your input, Karen Heckford St. Mary's Medical Center San Francisco, California From Sjohnso616 <@t> aol.com Wed Oct 26 13:50:29 2005 From: Sjohnso616 <@t> aol.com (Sjohnso616@aol.com) Date: Wed Oct 26 13:50:47 2005 Subject: [Histonet] another CAP question ANP21150 Message-ID: <1e5.47d7190c.309129f5@aol.com> Hi All, I need to ask any of you have dealt with this to share experiences. The question "Are slides identified adequately?". The note of explanation on the checklist for this question goes further to state "If slides are manually prelabeled, the original numbers on the glass must be readable from the underside of the slide" Our immunos are written by hand and a barcoded label is placed on the slide. We are using Fisher slides that are made for etching that have the darker coating on the back. Are we at risk for a defieciency? From azdudley <@t> hotmail.com Wed Oct 26 14:08:24 2005 From: azdudley <@t> hotmail.com (anita dudley) Date: Wed Oct 26 14:08:34 2005 Subject: [Histonet] job opening Message-ID: providence hospital in mobile alabama has a full time position in the histology dept. it would be performing all duties of histology, no frozen section our pathologists do their own. please contact wayne kreko at 251-366-1401 or dr. john millman 251-633-1407 if you are interested. anita dudley providence hosp. From bill501 <@t> mindspring.com Wed Oct 26 14:15:06 2005 From: bill501 <@t> mindspring.com (Bill B.) Date: Wed Oct 26 14:15:23 2005 Subject: [Histonet] Expired Stains In-Reply-To: References: Message-ID: At 10:57 AM -0700 10/26/05, Heckford, Karen - SMMC-SF wrote: >Hi Everyone, I was wondering if anyone knows how JCAHO views expired >Special Stains. I have some expired Special Stains. We do low volume, >trying to use up some stains before they expire can be difficult. In the name of controlling medical costs... We verify the stains by performing them on standard controls of tissues they might be used for and document the findings. We place a sticker with a new exp date for 6 months on the bottles and jars. Of course every stain has a control on the same slide which provides another verification. We are not JCAHO. I have jars of methylene blue and gold chloride powder from the '50's which work beautifully today, not that we use them much. I keep them with my antique collection so the inspectors dont bitch about expired reagents. In the early 80's I made my own anti-GFAP from human spinal cords, before it was commercially available. It was put in microtubes stored in a Revco. It is still giving strong specific staining. I am not using this on human material, though it would work fine. I am sure the manufacturers are strongly against this and lobby the feds heavily. BB From Matthew_Frank <@t> URMC.Rochester.edu Wed Oct 26 14:32:55 2005 From: Matthew_Frank <@t> URMC.Rochester.edu (Frank, Matthew) Date: Wed Oct 26 14:33:15 2005 Subject: [Histonet] Expired Stains Message-ID: The Biological Stain Commission has performed a stability study on dye powders and found that dye powders kept in closed containers stained well after over 30 years of storage. Solutions of course are a different story. -----Original Message----- From: Heckford, Karen - SMMC-SF [mailto:Karen.Heckford@CHW.edu] Sent: Wednesday, October 26, 2005 1:57 PM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] Expired Stains Hi Everyone, I was wondering if anyone knows how JCAHO views expired Special Stains. I have some expired Special Stains. We do low volume, trying to use up some stains before they expire can be difficult. I can still get the stains to work and the Pathologists like them. Should I throw them away and order new? As you know the stains can be costly. Thanks for your input, Karen Heckford St. Mary's Medical Center San Francisco, California _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From osteffe <@t> online.no Wed Oct 26 14:38:09 2005 From: osteffe <@t> online.no (ole) Date: Wed Oct 26 14:38:20 2005 Subject: [Histonet] Anti human - alpha 3,4,5 or 6 type iv collagen Message-ID: <000a01c5da64$cbd02dd0$0100000a@ole> Hi Im looking for anti-human antibodies against; Alpha (3, 4, 5 or 6) type IV Collagen. (Alport syndrome) Do anyone have recommandations of some of these antibodies? I would prefer that they works on formalinfixed tissue, but we have also fresh-frozen and alcohol fixed tissue from this kidney. Best regards Ole Johnny Steffensen Dep of Pathology Aalesund Norway From LuckG <@t> empirehealth.org Wed Oct 26 16:34:53 2005 From: LuckG <@t> empirehealth.org (Luck, Greg D.) Date: Wed Oct 26 16:35:08 2005 Subject: [Histonet] CAP question on grossing Message-ID: Charles (et.al.), Where "specifically" is it stated in any regulations (e.g. CLIA '88, CAP, Federal Register or other source) that "tissue grossing" is defined as high complexity testing? Thanks, Greg Greg Luck, BS, HT(ASCP) Anatomic Pathology Supervisor Deaconess Medical Center 800 W. 5th Ave Spokane, WA 99204 Phone 509.473.7077 Fax 509.473.7133 luckg@empirehealth.org www.deaconessmedicalcenter.org -----Original Message----- From: Charles.Embrey [mailto:Charles.Embrey@carle.com] Sent: Wednesday, October 26, 2005 7:49 AM To: Jesus Ellin Cc: Histonet@pathology.swmed.edu Subject: RE: [Histonet] CAP question on grossing CAP and CLIA have never made a separation in "transference" from grossing. Ever CLIA statement I have ever read on the subject states that grossing any tissue is high complexity testing. "Transference" is not even recognized in the CAP checklist. I routinely assist on CAP inspections and I would love to see a lab try to sell me on "transference". Of course it is only a Phase II write up but if the problem is not documented as corrected the lab can lose its accreditation. Liability and appropriate pay are other considerations I personally have mentioned in other e-mails but have been flamed for. My personal feeling is that if someone wants to undervalue their talent and work that is up to them. As long as labs can get techs to do part of the pathologists job for about a tenth of the pathologist's salary, those people will be prayed upon. Charles Embrey, PA(ASCP) -----Original Message----- From: Jesus Ellin [mailto:JEllin@yumaregional.org] Sent: Tuesday, October 25, 2005 7:36 PM To: Charles.Embrey Subject: RE: [Histonet] CAP question on grossing What about transference, our pathologist's are saying that small bx's or anything that does not require cutting or manipulation falls under this category. They are saying any registered or registry eligable can do this portion of the gross. What needs to happen is have stated black in white what techs can or can not do. This not only puts a lot of liability on the tech, it also is having tech do a PA or pathologist job at a techs wage, which is not right at all. Were can someone find out what the scale is for grossing tech's? I know it depends on the institution, but there has to be some sort of support for this other than requirements that CLIA and CAP set forth. Also what other than a phase two what can also happen if a facility is caught doing this.?? Jesus Ellin Yuma Regional Medical Center _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From JMahoney <@t> alegent.org Wed Oct 26 17:10:04 2005 From: JMahoney <@t> alegent.org (Janice A Mahoney) Date: Wed Oct 26 17:10:35 2005 Subject: [Histonet] CAP question on grossing Message-ID: S493.1489(b) and S493.1449 (b), (l) or(m) of the Federal Register. You can get interpretive guidelines that help make this document easier to understand. As long as individuals qualify under S493.1489, the technical supervisor (also defined under CLIA88) may deligate gross examination. Gross examination is defined as the physical examination/description, including color, weight, measurement and other characteristics of the tissue; or other mechanical procedures for which specific written protocol has been developed. Hope this helps. Jan Mahoney Omaha, NE >>> "Luck, Greg D." 10/26/2005 4:34 PM >>> Charles (et.al.), Where "specifically" is it stated in any regulations (e.g. CLIA '88, CAP, Federal Register or other source) that "tissue grossing" is defined as high complexity testing? Thanks, Greg Greg Luck, BS, HT(ASCP) Anatomic Pathology Supervisor Deaconess Medical Center 800 W. 5th Ave Spokane, WA 99204 Phone 509.473.7077 Fax 509.473.7133 luckg@empirehealth.org www.deaconessmedicalcenter.org -----Original Message----- From: Charles.Embrey [mailto:Charles.Embrey@carle.com] Sent: Wednesday, October 26, 2005 7:49 AM To: Jesus Ellin Cc: Histonet@pathology.swmed.edu Subject: RE: [Histonet] CAP question on grossing CAP and CLIA have never made a separation in "transference" from grossing. Ever CLIA statement I have ever read on the subject states that grossing any tissue is high complexity testing. "Transference" is not even recognized in the CAP checklist. I routinely assist on CAP inspections and I would love to see a lab try to sell me on "transference". Of course it is only a Phase II write up but if the problem is not documented as corrected the lab can lose its accreditation. Liability and appropriate pay are other considerations I personally have mentioned in other e-mails but have been flamed for. My personal feeling is that if someone wants to undervalue their talent and work that is up to them. As long as labs can get techs to do part of the pathologists job for about a tenth of the pathologist's salary, those people will be prayed upon. Charles Embrey, PA(ASCP) -----Original Message----- From: Jesus Ellin [mailto:JEllin@yumaregional.org] Sent: Tuesday, October 25, 2005 7:36 PM To: Charles.Embrey Subject: RE: [Histonet] CAP question on grossing What about transference, our pathologist's are saying that small bx's or anything that does not require cutting or manipulation falls under this category. They are saying any registered or registry eligable can do this portion of the gross. What needs to happen is have stated black in white what techs can or can not do. This not only puts a lot of liability on the tech, it also is having tech do a PA or pathologist job at a techs wage, which is not right at all. Were can someone find out what the scale is for grossing tech's? I know it depends on the institution, but there has to be some sort of support for this other than requirements that CLIA and CAP set forth. Also what other than a phase two what can also happen if a facility is caught doing this.?? Jesus Ellin Yuma Regional Medical Center _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From JEllin <@t> yumaregional.org Wed Oct 26 19:53:57 2005 From: JEllin <@t> yumaregional.org (Jesus Ellin) Date: Wed Oct 26 19:54:26 2005 Subject: [Histonet] CAP question on grossing Message-ID: This is great it is nice to know that we are not the only hospital with this issue. This is something that the ASCP needs to address, and not only have the CLIA guideline. Currently there are a lot of places were tech are asked to gross, so if this is a High Complexity testing, why is this not in the test to become a Histo tech. Further more it seems that the hospital, research and private labs, not to mention the Dr's are benefitting from this. I have to agree with Charles that you need to be paid for the service that you are doing, but what happens when management does not want to pay or they say show me or give me information on were salaries reflect this. This brings us to several questions, were we need our state, national and peers to get together and figure out a solution. There are already a huge shortage of tech, not to mention Pathologist Assistants and Pathologist. I see the need to adapt to the environment that is being created, but not at the expense of the patient or quality of our work. Then why be a Pathologist Assistant if you can gross, do autopsies and be a histotech.?? There needs to be some sort of standard here. I know that I am getting excited, but the fact of the matter is, We PAY dues to these societies and in recent all they have done is collect. The time has come to get the ball rolling and get what we are due, whether it be titles, levels of competencies or pay, something need to be done. Jesus Ellin Yuma Regional Medical Center From kwuny <@t> email.cs.nsw.gov.au Wed Oct 26 21:41:18 2005 From: kwuny <@t> email.cs.nsw.gov.au (Young Kwun) Date: Wed Oct 26 21:39:42 2005 Subject: [Histonet] Anti human - alpha 3,4,5 or 6 type iv collagen In-Reply-To: <000a01c5da64$cbd02dd0$0100000a@ole> Message-ID: <200510271239558.SM01416@crgcsls814> Ole, You can get some of the antibodies from Kamiya Biomedical company. They have Collagen IV, alpha-1,3 & 5 monoclonal antibodies. Alpha 3 and 5 work on paraffin sections, I am still trying to improve the alpha-1 antibody staining. Try their website www.kamiyabiomedical.com 12779 Gateway Drive, Seattle, WA 98168, USA Tel: (206) 575-8068 Fax: (206) 575-8094 E-mail: webmaster@kamiyabiomedical.com Cheers, Young Kwun Senior Hospital Scientist Dept of Anatomical Pathology Concord Hospital Concord NSW 2139 Australia Phone:61-2-9767 6075 Fax:61-2-9767 8427 Email:kwuny@email.cs.nsw.gov.au -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of ole Sent: Thursday, 27 October 2005 5:38 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Anti human - alpha 3,4,5 or 6 type iv collagen Hi Im looking for anti-human antibodies against; Alpha (3, 4, 5 or 6) type IV Collagen. (Alport syndrome) Do anyone have recommandations of some of these antibodies? I would prefer that they works on formalinfixed tissue, but we have also fresh-frozen and alcohol fixed tissue from this kidney. Best regards Ole Johnny Steffensen Dep of Pathology Aalesund Norway _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _____________________________________________________________________ This e-mail has been scanned for viruses by MCI's Internet Managed Scanning Services - powered by MessageLabs. For further information visit http://www.mci.com "This message is intended for the addressee named and may contain confidential information. If you are not the intended recipient, please destroy it and notify the sender. Views expressed in this message are those of the individual sender, and are not necessarily the views of the Sydney South West Area Health Service." From anh2006 <@t> med.cornell.edu Wed Oct 26 23:13:00 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Wed Oct 26 23:08:09 2005 Subject: [Histonet] Type I and Type II pneumocytes Message-ID: I am looking for markers against Type I and Type II pneumocytes in human and mouse tissues for IHC. Any suggestions? Markers, clones, catalog #s, protocols - all are welcome. Thanks, Andrea -- From lpwenk <@t> sbcglobal.net Thu Oct 27 04:38:26 2005 From: lpwenk <@t> sbcglobal.net (Lee & Peggy Wenk) Date: Thu Oct 27 04:39:18 2005 Subject: [Histonet] CAP question on grossing In-Reply-To: Message-ID: <1081716711-75562105@pathology.swmed.edu> I can address, somewhat, the part about why is grossing not on the ASCP certification exams. >From what I have heard, it will be in the future. The reason it has not been on, is that that HT exam had 3 routes, one of which was the high school diploma + 1 year on-the-job-training (OJT) route. That route did not meet the CLIA criteria. >From what was said a couple of years ago (I believe by an ASCP rep at the Instructors of Histotechnology forum at an NSH Symposium), the plan was to wait until the HS route was dropped. That would leave the NAACLS route and the associate degree/1 year OJT routes. Those with the associate degree would meet the CLIA criteria. Maybe someone else on the ASCP Histotechnology Exam Committee can fill in more details. Peggy A. Wenk, HTL(ASCP)SLS William Beaumont Hospital Royal Oak, MI 48037 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Jesus Ellin Sent: Wednesday, October 26, 2005 8:54 PM To: JMahoney@alegent.org; Charles.Embrey@carle.com; LuckG@empirehealth.org Cc: Histonet@pathology.swmed.edu Subject: RE: [Histonet] CAP question on grossing This is great it is nice to know that we are not the only hospital with this issue. This is something that the ASCP needs to address, and not only have the CLIA guideline. Currently there are a lot of places were tech are asked to gross, so if this is a High Complexity testing, why is this not in the test to become a Histo tech. Further more it seems that the hospital, research and private labs, not to mention the Dr's are benefitting from this. I have to agree with Charles that you need to be paid for the service that you are doing, but what happens when management does not want to pay or they say show me or give me information on were salaries reflect this. This brings us to several questions, were we need our state, national and peers to get together and figure out a solution. There are already a huge shortage of tech, not to mention Pathologist Assistants and Pathologist. I see the need to adapt to the environment that is being created, but not at the expense of the patient or quality of our work. Then why be a Pathologist Assistant if you can gross, do autopsies and be a histotech.?? There needs to be some sort of standard here. I know that I am getting excited, but the fact of the matter is, We PAY dues to these societies and in recent all they have done is collect. The time has come to get the ball rolling and get what we are due, whether it be titles, levels of competencies or pay, something need to be done. Jesus Ellin Yuma Regional Medical Center _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Lance.Erickson <@t> ihc.com Thu Oct 27 08:28:15 2005 From: Lance.Erickson <@t> ihc.com (Lance Erickson) Date: Thu Oct 27 08:28:54 2005 Subject: [Histonet] CAP question on grossing Message-ID: <8B08EC394B366D4A895DD5686D6AFE4A094037@LP-EXCHVS08.CO.IHC.COM> See the new federal register CLIA intrepretive guidelines appendix C subpart M that are effective April 24, 2003. At www.cms.hhs.gov/clia/apcsubm.pdf under qualifications of high complexity testing personnel section 493.1489 (b)(7) it is clear that all tissue gross examination whether it is color, measurement, or advanced dissection is considered high complexity testing and individuals performing this type of testing must qualify under this section. That is why the new CAP question ANP 11610 was instated and is effective April 28, 2005. CAP must abide by CLIA regulations and CLIA is part of the Center for Medicare and Medicaid Services which is a section of the US government's department of Health and Human Services. So if you would like to maintain your CLIA license and CAP certification and be paid by Medicare you must abide by the requirements for high complexity testing personnel for each person performing any kind of gross examination. Lance Erickson Anatomic Pathology Supervisor Primary Children's Medical Center Salt Lake City, UT -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Luck, Greg D. Sent: Wednesday, October 26, 2005 3:35 PM To: 'Charles.Embrey'; Jesus Ellin Cc: Histonet@pathology.swmed.edu Subject: RE: [Histonet] CAP question on grossing Charles (et.al.), Where "specifically" is it stated in any regulations (e.g. CLIA '88, CAP, Federal Register or other source) that "tissue grossing" is defined as high complexity testing? Thanks, Greg Greg Luck, BS, HT(ASCP) Anatomic Pathology Supervisor Deaconess Medical Center 800 W. 5th Ave Spokane, WA 99204 Phone 509.473.7077 Fax 509.473.7133 luckg@empirehealth.org www.deaconessmedicalcenter.org -----Original Message----- From: Charles.Embrey [mailto:Charles.Embrey@carle.com] Sent: Wednesday, October 26, 2005 7:49 AM To: Jesus Ellin Cc: Histonet@pathology.swmed.edu Subject: RE: [Histonet] CAP question on grossing CAP and CLIA have never made a separation in "transference" from grossing. Ever CLIA statement I have ever read on the subject states that grossing any tissue is high complexity testing. "Transference" is not even recognized in the CAP checklist. I routinely assist on CAP inspections and I would love to see a lab try to sell me on "transference". Of course it is only a Phase II write up but if the problem is not documented as corrected the lab can lose its accreditation. Liability and appropriate pay are other considerations I personally have mentioned in other e-mails but have been flamed for. My personal feeling is that if someone wants to undervalue their talent and work that is up to them. As long as labs can get techs to do part of the pathologists job for about a tenth of the pathologist's salary, those people will be prayed upon. Charles Embrey, PA(ASCP) -----Original Message----- From: Jesus Ellin [mailto:JEllin@yumaregional.org] Sent: Tuesday, October 25, 2005 7:36 PM To: Charles.Embrey Subject: RE: [Histonet] CAP question on grossing What about transference, our pathologist's are saying that small bx's or anything that does not require cutting or manipulation falls under this category. They are saying any registered or registry eligable can do this portion of the gross. What needs to happen is have stated black in white what techs can or can not do. This not only puts a lot of liability on the tech, it also is having tech do a PA or pathologist job at a techs wage, which is not right at all. Were can someone find out what the scale is for grossing tech's? I know it depends on the institution, but there has to be some sort of support for this other than requirements that CLIA and CAP set forth. Also what other than a phase two what can also happen if a facility is caught doing this.?? Jesus Ellin Yuma Regional Medical Center _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From MVaughan4 <@t> ucok.edu Thu Oct 27 09:25:29 2005 From: MVaughan4 <@t> ucok.edu (MVaughan4@ucok.edu) Date: Thu Oct 27 09:26:08 2005 Subject: [Histonet] Spleen stain thanks In-Reply-To: Message-ID: Thanks for those who responded to my request. Suggestions included trying rose bengal, mercurochrome, erythrosin, and concentrations of 0.5% up to 2% eosin-Y, as well as less rinsing afterward. I think everyone suggested phloxine to be added to the eosin. I am currently using 0.1% Eosin-Y in 69.5% EtOH with 0.5% glacial acetic acid. I am going to try adding some phloxine and perhaps upping the conc of the eosin. Thanks and I will return the favor when possible. Mel Melville B. Vaughan, Ph. D. Assistant Professor Department of Biology University of Central Oklahoma Edmond, OK 73034 From settembr <@t> umdnj.edu Thu Oct 27 09:51:48 2005 From: settembr <@t> umdnj.edu (Dana Settembre) Date: Thu Oct 27 09:52:53 2005 Subject: [Histonet] another CAP question ANP21150 Message-ID: Hello, whoever you are. You can e-mail that question directly to CAP for a response. They are very good about responding and doing so rapidly. The e-mail address is accred@cap.org Dana Settembre Immunohistochemistry Lab University Hospital - UMDNJ Newark, NJ USA >>> 10/26/2005 2:50:29 PM >>> Hi All, I need to ask any of you have dealt with this to share experiences. The question "Are slides identified adequately?". The note of explanation on the checklist for this question goes further to state "If slides are manually prelabeled, the original numbers on the glass must be readable from the underside of the slide" Our immunos are written by hand and a barcoded label is placed on the slide. We are using Fisher slides that are made for etching that have the darker coating on the back. Are we at risk for a defieciency? _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From djemge <@t> aol.com Thu Oct 27 10:46:39 2005 From: djemge <@t> aol.com (djemge@aol.com) Date: Thu Oct 27 10:46:52 2005 Subject: [Histonet] Mouse Tissue Processing Problems Help! Message-ID: <8C7A924C1151232-17A0-CF12@MBLK-M16.sysops.aol.com> Please help! Most of the mouse tissue (liver, brain, embryo's, pancreas) is very dry at times almost glassy. I came from a clinical histology lab and recently accepted a position as the only histotech in a research lab. The main tissue in this lab is Bouins fixed gonadal mouse tissue rinsed with water for 3 days, then 50% ETOH for 3 days, and 4 to 8 hour BNF fixed mouse tissue that is transferred to 70% ETOH (For later IF and IHC) then given to me. The Bouins fixed mouse testes and epididymous seem fine although somewhat crystal like at the inside edges. Formalin fixed testes transferred to the 70% has no crystals but is dry. All parameters and protocols were established by my predecessor. Since I am not familiar with mouse tissue I want to get this right! Processor Set up on Sakura VIP 2000 Enclosed Processor Using Surgipath Reagent Alcohol 70% Alcohol 30 mins Retort temp 38 degees C P/V on (the tissue sits in this for delay) 70% Alcohol 30 mins " " 80% Alcohol 30 mins " " 95% Alcohol 30 mins " " 100% Alcohol 30 mins " " 100% Alcohol 30 mins " " 100% Alcohol 30mins " " Xylene 30mins " " Xylene 30mins " " Xylene 30mins " " Paraffin 30mins Temp set at 58 degrees C (although retort temp says 60 C) P/V on Paraffin 30mins " " Paraffin 30mins " " Help with Paraffin: The Paraffin was Surgipath Formula R, but this crackled when put on ice. I switched to Cardinal Health's Ameraffin Cat no. M7346-1A I changed the processor station temps and embedding station temps to 58 degrees C per manufactures instructions. I can't say I am satisfied with this paraffin either as it is sticky and I need a waterbath temp of 38 degrees to cut it or it spreads quickly (I used this at my old lab for human tissue, but it seems to respond differently here - I manually checked the temps). The Formula R had a waterbath temp of 47 degrees and my predecessor had the processor station temps at 60 C and the retort temp had 63 to 64 C indicated, and embedding center at 62C. From Eric <@t> ategra.com Wed Oct 26 20:21:28 2005 From: Eric <@t> ategra.com (Eric Dye (ext 223)) Date: Thu Oct 27 11:03:04 2005 Subject: [Histonet] Immediate opportunities for Histotechnologists Message-ID: Histonetters I'm presently on a search for my best client companies seeking HistoTech Supervisors and Histo Bench Techs. These are mostly fulltime permanent positions. (I have a few travel/temp HistoTech positions as well - see below). Most of my Histo Techs positions are clinical, although I do have a few Research Histo Tech positions as well. Here are some of my Hottest jobs: 1. Louisiana( North Western) Openings for a HistoTech(Bench) No weekends, No call, and Top Dollar. 2. Ohio(Dayton Area) HistoTech Manager No Weekends, No call, Top Dollar if you are interested, please call me today at 1-800-466-9919 ext 223 3. Colorado (Greater Denver area) Opening for Several Bench Positions(Night Shift) No weekends, No call, Top Dollar 4. Georgia Openings for Histotech(Temp to Perm) No Weekends, No Call, Top Dollar, Excellent Benefits if you are interested, please call me today at 1-800-466-9919 ext 223 5. Minnesota Opening for HistoTech(Bench) No weekends, No Call, Top Dollar 6. Pennsylvania(Philadelphia area) Openings for HistoTech(Bench) No Weekends, No Call, Top Dollar if you are interested, please call me today at 1-800-466-9919 ext 223 7.Rhode Island Openings for Histotech(Bench) No weekends, No call, Top Dollar 8. California( Southern) HistoTech Supervisor(2nd shift) Openings for Histotechs(Bench,2nd shift) Great Location, No weekends, No call, Top Dollar 9. California(Southern) Openings for HistoTechs(Bench) Great Location, No weekends, No call, Top Dollar 10. Pennsylvania(Multiple jobs in greater Pittsburgh area) HistoTech opening(Bench) if you are interested, please call me today at 1-800-466-9919 ext 223 11. Illinois(Multiple jobs in Greater Illinois area) Opening for Histotech(Bench) 12. Michigan(Multiple jobs in Greater Michigan area) Openings for Histotechs(Bench) 13. Indiana Opening for Histotech(Bench) 14. Washington State(Eastern) Opening for Lead HistoTech(Bench) No weekends, No call if you are interested, please call me today at 1-800-466-9919 ext 223 15. Oklahoma Opening for Histotech(Bench) No weekends, No call. 16. Massachusetts (Boston Area) Part time Histo Tech(Permanent) No weekends, No call 17. Tennesee(Memphis area) Openings for Supervisor and Bench Techs 18. New Mexico openings for Supervisor and Bench Techs No Weekends, No Call, Excellent Benefits.. 19. Nebraska Openings for Histo Techs(Bench) No Weekends, No Call, Top Dollar 20. Virginia(Western Virginia) Opening for Bench Histotech No weekends, No call if you are interested, please call me today at 1-800-466-9919 ext 223 21. Ohio( Southern) Opening for Bench HistoTech No weekends, No call 22. Maryland( Baltimore area) Opening for Bench Histotech 23. Missouri(Greater Missouri area) Opening for Bench Histotech No weekends, No call 24. Wisconsin(Eastern area) Opening for a Bench Histotech No weekends, No call 25. Florida(Greater Florida area) Openings for Bench Histotech No weekends, No call The clients are currently interviewing candidates - so if you are interested, please call me today at 1-800-466-9919 ext 223 Thank you, Eric Dye-Sr Allied Healthcare Recruiter 1-800-466-9919 ext 223 From weneng2004 <@t> yahoo.com Thu Oct 27 12:27:49 2005 From: weneng2004 <@t> yahoo.com (wen eng) Date: Thu Oct 27 12:28:04 2005 Subject: [Histonet] Decloaking Chamber (high pressure cooker) Message-ID: <20051027172749.97588.qmail@web53407.mail.yahoo.com> Hello, Does anybody use Decloaking Chamber from Biocare Medical? How should I set the SP1 temperature and time if I need 20 minutes treatment? The factory setting is 125 degree 30 seconds and the tech support said 20 minutes is too long. She suggested me to keep 30 seconds. I still want the suggestion from here. How do you set your program? Thanks! Wen --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. From gcallis <@t> montana.edu Thu Oct 27 12:30:02 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Thu Oct 27 12:30:15 2005 Subject: [Histonet] Help!! We have a problem with a Mounting Media Message-ID: <6.0.0.22.1.20051027111608.01b0de68@gemini.msu.montana.edu> Is anyone experiencing this problem with Protocol Mounting Medium T, toluene solvent based from Fisher. They take 50 mls and put it into a bottle with a secure, air tight, solvent resistant lid in order to not dip into stock bottle all the time. If they use the mounting media absolutely fresh after aliquoting, the mounting media drys nicely, everything is fine. They are coverslipping cytospin stained blood cells, Diff Quik stained, and dry without any immersion oil used before coverslipping. If they let the Protocol media aliquot, even though in an air tight, new bottle, sit around, they notice a layer or some type of separation within top layer of media. When they use this now separated aliquot, stirred, and then try to dry slide, the coverslip still slips and slides around - failure of drying!! Very messy! Something is happening, either evaporation of toluene to change ratio of solvent to acrylic resin, maybe something else i.e faulty manufacturing/preparation of this media? This is a total mystery to me as I have done this type of aliquoting with other mounting medias for years and NEVER had a problem except to add solvent to restore media - thick to thinner. Nor I have never experienced layering/separation phenomena with my other mounting media (Richard Allan brand or Permount). After coverslipping, the latter mounting media set up and dry nicely with no slip/sliding away!!! Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From kmerriam2003 <@t> yahoo.com Thu Oct 27 12:31:19 2005 From: kmerriam2003 <@t> yahoo.com (Kim Merriam) Date: Thu Oct 27 12:31:27 2005 Subject: [Histonet] mouse heart matrix Message-ID: <20051027173119.75498.qmail@web50310.mail.yahoo.com> Hello all, I was wondering if anyone knew of a vendor that sells a tissue matrix that will allow me to take thin cross-sections of mouse heart (something similar to the brain matrices sold by Ted Pella. Regards, Kim Kim Merriam Novartis Cambridge, MA --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. From Jackie.O'Connor <@t> abbott.com Thu Oct 27 12:34:37 2005 From: Jackie.O'Connor <@t> abbott.com (Jackie M O'Connor) Date: Thu Oct 27 12:35:04 2005 Subject: [Histonet] Decloaking Chamber (high pressure cooker) Message-ID: I'm using the factory settings for virtually all antibodies - so far, I haven't found the need to change the settings for anything. I did uncover a problem, however, with the computer chip in my old Decloaker - the temperature and pressure were not reaching the set point - but I didn't notice until I sat down to actually watch the machine. The electronic temperature display was bouncing all over the place. I had to get a new machine. Jackie O' wen eng Sent by: histonet-bounces@lists.utsouthwestern.edu 10/27/2005 12:27 PM To: histonet cc: (bcc: Jackie M O'Connor/LAKE/GPRD/ABBOTT) Subject: [Histonet] Decloaking Chamber (high pressure cooker) Hello, Does anybody use Decloaking Chamber from Biocare Medical? How should I set the SP1 temperature and time if I need 20 minutes treatment? The factory setting is 125 degree 30 seconds and the tech support said 20 minutes is too long. She suggested me to keep 30 seconds. I still want the suggestion from here. How do you set your program? Thanks! Wen --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From la.sebree <@t> hosp.wisc.edu Thu Oct 27 12:43:23 2005 From: la.sebree <@t> hosp.wisc.edu (Sebree Linda A.) Date: Thu Oct 27 12:43:40 2005 Subject: [Histonet] Decloaking Chamber (high pressure cooker) Message-ID: We translated 20 minutes HIER in a water bath to around 2-3 minutes in the Decloaking Chamber. Hope this helps. Linda A. Sebree University of Wisconsin Hospital & Clinics IHC/ISH Clinical & Research Laboratory DM223-VA 600 Highland Ave. Madison, WI 53792 (608)265-6596 FAX: (608)262-7174 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of wen eng Sent: Thursday, October 27, 2005 12:28 PM To: histonet Subject: [Histonet] Decloaking Chamber (high pressure cooker) Hello, Does anybody use Decloaking Chamber from Biocare Medical? How should I set the SP1 temperature and time if I need 20 minutes treatment? The factory setting is 125 degree 30 seconds and the tech support said 20 minutes is too long. She suggested me to keep 30 seconds. I still want the suggestion from here. How do you set your program? Thanks! Wen --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From shive003 <@t> umn.edu Thu Oct 27 13:01:35 2005 From: shive003 <@t> umn.edu (Jan Shivers) Date: Thu Oct 27 13:01:44 2005 Subject: [Histonet] Decloaking Chamber (high pressure cooker) References: <20051027172749.97588.qmail@web53407.mail.yahoo.com> Message-ID: <005901c5db20$78b82e60$41065486@auxs.umn.edu> I use a Decloaking Chamber for Chronic Wasting Disease slides, and yes, 20 minutes is the correct setting for that particular test, with a 25 minute cooldown. Most other IHC tests don't require a lengthy setting on the Decloakers. I'd recommend contacting Biocare to do this for you. They came and installed the settings on my chambers. Jan Shivers U of MN Vet Diag Lab ----- Original Message ----- From: "wen eng" To: "histonet" Sent: Thursday, October 27, 2005 12:27 PM Subject: [Histonet] Decloaking Chamber (high pressure cooker) > Hello, > > Does anybody use Decloaking Chamber from Biocare Medical? How should I set the SP1 temperature and time if I need 20 minutes treatment? The factory setting is 125 degree 30 seconds and the tech support said 20 minutes is too long. She suggested me to keep 30 seconds. I still want the suggestion from here. How do you set your program? > > Thanks! > Wen > > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > From TJJ <@t> Stowers-Institute.org Thu Oct 27 14:24:14 2005 From: TJJ <@t> Stowers-Institute.org (Johnson, Teri) Date: Thu Oct 27 14:24:44 2005 Subject: [Histonet] Re: Red HRP chromagen/substrate Message-ID: Andrea, It depends on what you consider "best". As Gayle pointed out, for some it's endpoint color. For others, it's permanence. AEC needs to be mounted in aqueous mountant because it's not stable in organic solvents (even the polyvinyl alcohol in some of the aqueous mountants can cause fading!). The way to get around this is to either use a permanent chromogen (Vector Nova Red), or you can use Crystal mount to cover the AEC stained tissue sample and allow to dry. We always coverslip it with permanent mountant for better visualization under the microscope. Teri Johnson Managing Director Histology Facility Stowers Institute for Medical Research 1000 E. 50th St. Kansas City, MO 64133 From jkiernan <@t> uwo.ca Thu Oct 27 14:36:27 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Thu Oct 27 14:36:33 2005 Subject: [Histonet] Cholesterol staining suggestions References: <6.0.0.22.1.20051026114957.01b4d488@gemini.msu.montana.edu> Message-ID: <43612C3B.BD853396@uwo.ca> The only specific method for cholesterol that doesn't involve a nasty acid is the cholesterol oxidase-DAB method (Emeis et al 1977 Histochem. J. 9:197-204). It can also detect cholesterol esters if the sections are first treated with cholesterol esterase. It's rather more than a "stain" but should be OK for research. It can also be used at the EM level (eg Pelletier & Vitale 1994 J Histochem Cytochem 42:1539-1554.) Regarding the traditional stains for cholesterol: According to Adams, Pearse and Bayliss High, the perchloric acid-naphthoquinone method is the most specific and sensitive one for cholesterol and its esters. I've tried it once or twice; it works but it's rather rough on the sections. The blue colour fades after a few hours. An older, simpler method is that of Schultz (here summarized from Bayliss High's 1984 "Lipid Histochemistry" p.40-41). I've not tried it myself. Frozen sections, after formal-calcium fixation, air-dried onto slides. 1. Oxidize sections in 2.5% iron alum in 0.2M acetate buffer, pH 3 for 4 hours at 37C. 2. Wash, 3 X 20 min in the acetate buffer. 3. Rinse in distilled water. 4. Treat with 5% formalin in tap water (ie 2% formaldehyde), 10 min. 5. Drain and allow to dry. 6. Apply a drop or two of Schulz reagent (see below), and then a coverslip. Examine with microscope. Result. Cholesterol blue-green, fading quickly. Schulz reagent: Add 1ml glacial acetic acid dropwise to 1 ml conc. sulphuric acid, with agitation. It becomes hot and syrupy; let it cool. Cholesterol and its esters are birefringent with crossed polars. This property was discovered by B. Doinikow (1913) Deutsche Z. Nervenheilk. 46:20-42) and it has been exploited to trace degenerating myelinated axons in frozen sections of human brain (Miklossy & Van der Loos 1987 Brain Res. 426:377-380). I tried to reproduce their results in the 1990s but the sections contained great quantities of birefringent dirt (completely invisible with ordinary illumination). The dirt didn't consist of tiny needle-like crystals like the ones described by Doinikow and shown in the colour photos in Miklossy's paper. My material had been stored for a long time in formalin before cutting, so perhaps lipids everywhere had changed into some sort of birefringent junk. -- ------------------------------- John A. Kiernan Department of Anatomy and Cell Biology The University of Western Ontario London, Canada N6A 5C1 kiernan[AT]uwo.ca http://publish.uwo.ca/~jkiernan/ http://instruct.uwo.ca/anatomy/530/index.htm _______________________________ Gayle Callis wrote: > > I had an inquiry about what staining method is best for cholesterol but > after looking at the methods (PAN) in the literature I have on hand, I am > not sure I want to deal with this type of staining protocol. Would it be > better to do an immunostain? If so, this would be on murine tissue. > > Any suggestions are welcome for the grad student asking about cholesterol > > Gayle Callis > Research Histopathology Supervisor > Veterinary Molecular Biology > Montana State University - Bozeman > PO Box 173610 > Bozeman MT 59717-3610 > 406 994-6367 > 406 994-4303 (FAX) > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rocan <@t> mac.com Thu Oct 27 14:39:52 2005 From: rocan <@t> mac.com (Rocan) Date: Thu Oct 27 14:40:14 2005 Subject: [Histonet] mouse heart matrix In-Reply-To: <20051027173119.75498.qmail@web50310.mail.yahoo.com> References: <20051027173119.75498.qmail@web50310.mail.yahoo.com> Message-ID: <85AEC9B5-EF4C-42BF-A6BE-04EAA324F828@mac.com> Yes, There are these matrices for heart We have just bought these matrices from them here is their web address: http://www.zivic-miller.com/ Good Luck, Rocio ----- Dr.Rocio Sierra-Honigmann Director Engineered Wound Repair Laboratory Cedars Sinai Research Institute Davis 1091 310-423-1882 Honigmannr@cshs.org On Oct 27, 2005, at 10:31 AM, Kim Merriam wrote: > Hello all, > > I was wondering if anyone knew of a vendor that sells a tissue > matrix that will allow me to take thin cross-sections of mouse > heart (something similar to the brain matrices sold by Ted Pella. > > Regards, > Kim > > > Kim Merriam > Novartis > Cambridge, MA > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > From cfavara <@t> niaid.nih.gov Thu Oct 27 15:24:21 2005 From: cfavara <@t> niaid.nih.gov (Favara, Cynthia (NIH/NIAID)) Date: Thu Oct 27 15:24:30 2005 Subject: [Histonet] Decloaking Chamber (high pressure cooker) Message-ID: I use this almost exclusively as I work with prions as well. If you are working with new antibodies in a research setting I would experiment with the times. I think the company is giving guidelines for clinical applications. I have some antibodies that differ somewhat however in my experience 30seconds - 5 minutes @120C is adequate for most. I have found the company to be most helpful with adjusting the settings if necessary. c Cynthia Favara NIAID/NIH/RML/LPVD 903 South 4th Street Hamilton, MT 59840 406-363-9317 Disclaimer: The information in this e-mail and any of its attachments is confidential and may contain sensitive information. It should not be used by anyone who is not the original intended recipient. If you have received this e-mail in error please inform the sender and delete it from your mailbox or any other storage devices. National Institute of Allergy and Infectious Diseases shall not accept liability for any statements made that are sender's own and not expressly made on behalf of the NIAID by one of its representatives -----Original Message----- From: wen eng [mailto:weneng2004@yahoo.com] Sent: Thursday, October 27, 2005 10:28 AM To: histonet Subject: [Histonet] Decloaking Chamber (high pressure cooker) Hello, Does anybody use Decloaking Chamber from Biocare Medical? How should I set the SP1 temperature and time if I need 20 minutes treatment? The factory setting is 125 degree 30 seconds and the tech support said 20 minutes is too long. She suggested me to keep 30 seconds. I still want the suggestion from here. How do you set your program? Thanks! Wen --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From dharclerode <@t> cytoritx.com Thu Oct 27 15:40:07 2005 From: dharclerode <@t> cytoritx.com (Donna Harclerode) Date: Thu Oct 27 15:40:24 2005 Subject: [Histonet] Mouse Tissue Processing Problems Help Message-ID: <3DE0F644E093DF4BAE80C254176696A50204E0@mp-mailserver.macropore.com> My first suggestion is turn off the heat in your processor. I personally do not like tissues stored in 70% alcohol but others think it is great. Having heat on in 70% alcohol and your other solvents would be very drying to most tissue. Clinical labs use heat because they are pressed for time (not optimal, but it works). I recommend never having heat on a processor with animal tissue. I also suggest using a xylene substitute and not xylene. I like the aliphatic hydrocarbon substitutes but the limonene (orange smell that some people get sensitized to) also more gentle than xylene. I use a 1 hr schedule for most of my tissues- I use Anatech's ProPar (aliphatic hydrocarcarbon xylene substitute) and Prosoft (alcohol combination) on a VIP Processor (with pressure and vacuum but no heat). All my tissues are well fixed before I put them on the machine. I got into the habit of washing tissues when I was using the Zinc formalin and regular formalin tissues to keep the phosphate crystals out of the tissues. It also keeps the formalin out of my alcohols and keeps the processor lines cleaner. Anatech site is http://www.anatechltdusa.com/index.html They make great products and have the best technical support! Graded alcohols will work (I have used ethanol only for mice and isopropyl for rats, dogs and monkeys in the past), but I find the ProSoft to be easier and not as drying as any of the plain alcohols. I do not make dilutions of ProSoft, but just rotate the containers per Anatech recommendations. I still face off all my blocks and put them back on ice to rehydrate them a bit before I section them. When I do chrondocyte pellets, I drop the time in each station to 20 minutes since they are so tiny. I currently use the Surgipath embedding and processing paraffin but will be switching to Richard Allan Type 3 for processing and Type 9 for embedding from VWR or elsewhere. 1. NBF hold till process 2. tap water 30 min 3. ProSoft 1hr 4. ProSoft 1 hr 5. ProSoft 1 hr 6. ProSoft 1 hr 7. ProPar 1 hr 8. ProPar 1 hr 9. ProPar 1 hr 10. Wax 1hr 11. Wax 1 hr 12. Wax 1 hr Good luck Donna Harclerode, HT, (ASCP), HTL, QIHC Immunohistochemist Cytori Therapeutics 6740 Top Gun St. San Diego, CA 92121 858-458-0900 ext 322 dharclerode@cytoritx.com From niloof <@t> yahoo.com Thu Oct 27 15:49:21 2005 From: niloof <@t> yahoo.com (Niloufar Fozouni) Date: Thu Oct 27 15:49:35 2005 Subject: [Histonet] collagen embeding Message-ID: <20051027204922.88497.qmail@web35614.mail.mud.yahoo.com> Hello every one, I need to embed pure collagen in particular substances in different concentration and stain them (van gieson's method). I've tried embedding collagen in gel and gelatin as a mold and then embedding that in OCT for cryosectioning but, couldn't get uniform section and also after staining the collagen washed off. I need to do that as a calibration in polarized light microscope. If any one has suggestions please let me know. Thank you. Niloo Physics Dept. Oakland University __________________________________ Yahoo! FareChase: Search multiple travel sites in one click. http://farechase.yahoo.com From PBugelsk <@t> CNTUS.JNJ.COM Thu Oct 27 15:52:23 2005 From: PBugelsk <@t> CNTUS.JNJ.COM (Bugelski, Peter [CNTUS]) Date: Thu Oct 27 15:52:34 2005 Subject: FW: [Histonet] mouse heart matrix Message-ID: <7BF70FA941B9AE4783EAAF733762F1B505198260@CNTUSMAEXS3.na.jnj.com> You can get stainless steel mouse heart matrixes from Harvard Apparatus or ASI instruments. From failm <@t> musc.edu Thu Oct 27 16:26:19 2005 From: failm <@t> musc.edu (Mildred Fail) Date: Thu Oct 27 16:26:45 2005 Subject: [Histonet] questions about rhodanine-copper stain Message-ID: i keep the rhodanine stock a year. Our protocol is very simple Rena fail RHODANINE SATURATED SOLUTION (STOCK) Absolute Alcohol 100.0 ml 5 p-dimethylaminobenzylidene* 0.2 gm RHODANINE SOLUTION (WORKING) Rhodanine Saturated Solution 3.0 ml Distilled Water 47.0 ml Shake stock before measuring and mixing solutions. 1. Deparaffinize and hydrate to distilled water. 2. Using plastic Coplin Jars; preheat the Rhodanine solution in 1200 watt microwave on half power 30 seconds then incubate for 5 minutes. 3. Wash well in several changes of distilled water. 4. Mayer's Hematoxylin for 1 minute. 5. Rinse with distilled water. 6. A quick rinse in 0.5% Sodium Borax. 7. Rinse well with distilled water 8. Deparaffinize, clear, and mount It will fade if not cover slipped right away udrun Lang" 10/26/05 08:23AM >>> I am dealing with the rhodanine stain, because we shall establish it in our histolab. I've read three different protocols (AFIP, Churukian, webpath). AFIP and Churukian require sodiumacetat solutions to mix with the stocksolution, the others require just the dilution of the stocksolution in Aqua dest. Why? Do both versions work? Churukian demands coverslipping with aqueous medium, the others don't. Is the staining result soluble in ethanol? The stability of the 0,2% rhodanine/ethanol stocksolution is described between one day and three months. What is the right shelf live? I hope someone can explain it to me. Gudrun Lang General hospital Linz, Austria _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From weneng2004 <@t> yahoo.com Thu Oct 27 17:57:01 2005 From: weneng2004 <@t> yahoo.com (wen eng) Date: Thu Oct 27 17:57:10 2005 Subject: [Histonet] Thanks for your help! Message-ID: <20051027225701.49819.qmail@web53415.mail.yahoo.com> Thank you all who helped me with setting program for decloaking chamber! So far what I learned is that the factory setting works for most antibodies but few antibodies do need adjust time. Thanks again for your help! Wen --------------------------------- Yahoo! FareChase - Search multiple travel sites in one click. From gcallis <@t> montana.edu Thu Oct 27 18:08:17 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Thu Oct 27 18:08:29 2005 Subject: [Histonet] Help!! We have a problem with a Mounting Media In-Reply-To: References: Message-ID: <6.0.0.22.1.20051027170421.01b66168@gemini.msu.montana.edu> Dear Denise, It was a brown glass reagent bottle with a seal in lid said to be resistent to solvents. Interestingly there was one mounting media are put into white plastic bottle with the little pull up tip for dispensing, and this media worked great even out of this white plastic dispenser. It was sold for years by Fisher but it could really dispense a gigantic drop or wad of media if not careful. Gayle Callis At 02:17 PM 10/27/2005, you wrote: >Hi Gayle, >You didn't mention the type of bottle used...glass or some form of >plastic. >My thought is, if it's plastic, perhaps some sort of chemical reaction >is occurring between the media and plastic or that the toluene is >evaporating through the plastic, giving you the situation you speculated >on in your query. >Have a good day, >Denise > >-----Original Message----- >From: histonet-bounces@lists.utsouthwestern.edu >[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Gayle >Callis >Sent: Thursday, October 27, 2005 1:30 PM >To: Histonet@lists.utsouthwestern.edu >Subject: [Histonet] Help!! We have a problem with a Mounting Media > >Is anyone experiencing this problem with Protocol Mounting Medium T, >toluene solvent based from Fisher. > >They take 50 mls and put it into a bottle with a secure, air tight, >solvent >resistant lid in order to not dip into stock bottle all the time. >If they use the mounting media absolutely fresh after aliquoting, the >mounting media drys nicely, everything is fine. They are coverslipping >cytospin stained blood cells, Diff Quik stained, and dry without any >immersion oil used before coverslipping. > >If they let the Protocol media aliquot, even though in an air tight, new > >bottle, sit around, they notice a layer or some type of separation >within >top layer of media. When they use this now separated aliquot, stirred, > >and then try to dry slide, the coverslip still slips and slides around >- >failure of drying!! Very messy! > >Something is happening, either evaporation of toluene to change ratio of > >solvent to acrylic resin, maybe something else i.e faulty >manufacturing/preparation of this media? This is a total mystery to me >as >I have done this type of aliquoting with other mounting medias for years > >and NEVER had a problem except to add solvent to restore media - thick >to >thinner. Nor I have never experienced layering/separation phenomena >with >my other mounting media (Richard Allan brand or Permount). After >coverslipping, the latter mounting media set up and dry nicely with no >slip/sliding away!!! > > > >Gayle Callis >Research Histopathology Supervisor >Veterinary Molecular Biology >Montana State University - Bozeman >PO Box 173610 >Bozeman MT 59717-3610 >406 994-6367 >406 994-4303 (FAX) > > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet From rod.coombe <@t> imvs.sa.gov.au Thu Oct 27 19:12:30 2005 From: rod.coombe <@t> imvs.sa.gov.au (Rod Coombe) Date: Thu Oct 27 19:12:42 2005 Subject: [Histonet] Solvent Recyclers Message-ID: <000501c5db54$49652b70$d86d140a@ITP36079> I am interested in comments from people who have used solvent recylers. Is the recylced xylene suitable to use in processing machines? Are you recycling alcohol from the processing machines and have you had problems with xylene contamination in the alcohol through the processing machine? How long does it take to recycle 20 litres of alcohol? Is anybody recycling formalin in preference to having it commercially disposed of and if so how long does it take to recycle 20 litres of formalin? Which are the best recyclers? Thankyou Rod Coombe Manager Tissue Pathology Institute of Medical and Veterinary Science Frome Road, Adelaide, South Australia Phone +61 8 8222 3201 Fax +61 8 82223204 From anh2006 <@t> med.cornell.edu Thu Oct 27 19:25:42 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Thu Oct 27 19:21:00 2005 Subject: [Histonet] VEGF receptor antibodies Message-ID: I am curious as to what antibodies people are using for VEGF receptors (1, 2, or 3) IHC in human tissue (paraffin or frozen)? Any information you can provide is most appreciated. Thanks, Andrea -- From Jodiputnam <@t> aol.com Thu Oct 27 19:31:08 2005 From: Jodiputnam <@t> aol.com (Jodiputnam@aol.com) Date: Thu Oct 27 19:31:28 2005 Subject: [Histonet] Moh's information Message-ID: <218.c9880e5.3092cb4c@aol.com> Hello. I have been working in histology for close to 15 years now and recently got a part time job as a Mohs tech. I really like it and want to get as much information as possible about the procedure. Anyone working as a Mohs tech that would be willing to share tips and insight would be greatly appreciated. I've been looking on the net for Mohs related literature and so far the books have all been way out of my price range. Any information will be greatly appreciated. Thanks in advance!!!! Jodi From petepath <@t> yahoo.com Fri Oct 28 06:23:21 2005 From: petepath <@t> yahoo.com (Stephen Peters M.D.) Date: Fri Oct 28 06:23:30 2005 Subject: [Histonet] Moh's information Message-ID: <20051028112321.47790.qmail@web30402.mail.mud.yahoo.com> Jody, You might want to try my embedding toys. I have a number of very happy Mohs users. http://pathologyinnovations.com/index.html Stephen Stephen Peters M.D. Vice Chairman of Pathology Hackensack University Medical Center 201 996 4836 Pathology Innovations, LLC 410 Old Mill Lane, Wyckoff, NJ 07481 201 847 7600 www.pathologyinnovations.com From jkiernan <@t> uwo.ca Fri Oct 28 08:01:41 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Fri Oct 28 08:01:43 2005 Subject: [Histonet] Moh's information References: <218.c9880e5.3092cb4c@aol.com> Message-ID: <43622135.138B5E44@uwo.ca> A search for "Moh's" on www.hstosearch.com brings up about 700 hits! Many are ads for jobs, and many are explanations and technical tips. John Kiernan London, Canada. ----------------------------------------------------- Jodiputnam@aol.com wrote: > > Hello. I have been working in histology for close to 15 years now and > recently got a part time job as a Mohs tech. I really like it and want to get as > much information as possible about the procedure. Anyone working as a Mohs tech > that would be willing to share tips and insight would be greatly appreciated. > I've been looking on the net for Mohs related literature and so far the > books have all been way out of my price range. Any information will be greatly > appreciated. Thanks in advance!!!! > > > Jodi > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jkiernan <@t> uwo.ca Fri Oct 28 08:01:57 2005 From: jkiernan <@t> uwo.ca (John A. Kiernan) Date: Fri Oct 28 08:02:01 2005 Subject: [Histonet] collagen embeding References: <20051027204922.88497.qmail@web35614.mail.mud.yahoo.com> Message-ID: <43622145.AAE64D07@uwo.ca> Agar might be better than gelatin. Gelatin is made by boiling collagen, and it stains with the Van Gieson method. Van Gieson staining does not increase the birefringence of collagen fibres. The best staining method for this purpose is sirius red F3B in saturated aqueous picric acid. See Junqueira et al (1979) Histochemical Journal 11:447-455 for a thorough study of the technique. If measurements (optical; orientation etc) are to be made, it can be advantageous to reduce the concentration of sirius red F3B in the solution; see Canham et al (1991) Neurological Research 21:618-626. John Kiernan Anatomy, UWO London, Canada. ____________________________________________________ Niloufar Fozouni wrote: > > Hello every one, > I need to embed pure collagen in particular substances > in different concentration and stain them (van > gieson's method). > I've tried embedding collagen in gel and gelatin as a > mold and then embedding that in OCT for > cryosectioning but, couldn't get uniform section and > also after staining the collagen washed off. > I need to do that as a calibration in polarized light > microscope. > If any one has suggestions please let me know. > Thank you. > Niloo > Physics Dept. > Oakland University > > > __________________________________ > Yahoo! FareChase: Search multiple travel sites in one click. > http://farechase.yahoo.com > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From brett_connolly <@t> merck.com Fri Oct 28 09:35:29 2005 From: brett_connolly <@t> merck.com (Connolly, Brett M) Date: Fri Oct 28 09:36:09 2005 Subject: [Histonet] IHC Punctate staining in colon epithelium, nonspecific? Message-ID: <355C35514FEAC9458F75947F5270974D0796A8@usctmx1103.merck.com> Has anyone ever seen apparent non-specific, punctate staining in colon epithelium...looks to be in the Golgi area. It's coming up with both biotin and non-biotin based IHC. Brett Brett M. Connolly, Ph.D. Merck & Co., Inc. MRL, Imaging Research WP-44K PO Box 4 West Point, PA 19486 PH 215-652-2501 fax. 215-993-6803 e-mail. brett_connolly@merck.com ------------------------------------------------------------------------------ Notice: This e-mail message, together with any attachments, contains information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates (which may be known outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD and in Japan, as Banyu) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system. ------------------------------------------------------------------------------ From c.ingles <@t> hosp.wisc.edu Fri Oct 28 10:25:40 2005 From: c.ingles <@t> hosp.wisc.edu (Ingles Claire) Date: Fri Oct 28 10:25:52 2005 Subject: [Histonet] Moh's information Message-ID: <583D3E9A1E843445BD54E461D1A2F6F3025ACE66@uwhis-xchng2.hosp.wisc.edu> You lucky person you! I have been a Histotech for about 4 years and a Mohs tech just over a year. It took me about a week to adjust to cutting Mohs so don't get too worried. One of the biggest secrets I can share is Liquid Nitrogen. It allows the fat to be cut beautifully without the other tissue areas and mounting media to become overhard and friable as when an isopentane bath is used. Another biggie is to make sure you have the entire epidermal and deep surface frozen flat. I usually freeze my sections flat on the pressure plate then cover with media as quickly as possible. If you wait too long and the epidermal edge starts to turn white, the media will tend not to support the edge as well, and it has a greater tendency to chip out. Big No-No. If you need anything else in the future, let me know. I'll help out any way I can. Claire Ingles, HTL UW Hospitals & Clinics Mohs Clinic Lab Madison WI -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu on behalf of Jodiputnam@aol.com Sent: Thu 10/27/2005 7:31 PM To: histonet@lists.utsouthwestern.edu Cc: Subject: [Histonet] Moh's information Hello. I have been working in histology for close to 15 years now and recently got a part time job as a Mohs tech. I really like it and want to get as much information as possible about the procedure. Anyone working as a Mohs tech that would be willing to share tips and insight would be greatly appreciated. I've been looking on the net for Mohs related literature and so far the books have all been way out of my price range. Any information will be greatly appreciated. Thanks in advance!!!! Jodi _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From Tbarnhart <@t> primecare.org Fri Oct 28 10:31:14 2005 From: Tbarnhart <@t> primecare.org (Barnhart, Tammy) Date: Fri Oct 28 10:33:49 2005 Subject: [Histonet] RNase contamination during in-situ Message-ID: <1779904B5E82D511914C00D0B793339205BFDA6F@exchangent> I need some help...we are going to try using the Dako PNA ISH kit for Kappa/Lambda and are concerned about the RNase contamination issue. Can anyone give me an idea as to what product to use to take care of this on surfaces, glassware and reagents? Any and all suggestions would be appreciated. Tammy Barnhart, BS, HTL(ASCP) Anatomic Pathology Supervisor St. Alexius Medical Center Bismarck, ND Confidentiality Notice:This e-mail message is for sole use of intended recipient(s) and may contain confidential and privileged information. Any unauthorized review, use, disclosure, distribution, or copying is prohibited. If you are not the intended recipient, please contact the sender by replying to this e-mail and destroy/delete all copies of this e-mail message. From gisela <@t> vetmed.wsu.edu Fri Oct 28 11:21:33 2005 From: gisela <@t> vetmed.wsu.edu (Bailey, Gisela) Date: Fri Oct 28 11:21:39 2005 Subject: [Histonet] New Position Message-ID: <35CF12B690D8CA4E95375A36B4E7B44C0B68E8@cvm36.vetmed.wsu.edu> Search # 4139 ADVERTISEMENT COPY Tissue Processing Laboratory Manager Washington Animal Disease Diagnostic Laboratory College of Veterinary Medicine, Pullman, WA The histology laboratory in the Washington Disease Diagnostic Laboratory (WADDL) is currently seeking applicants for the full-time position of tissue processing laboratory manager. Responsibilities of the position include providing management and leadership direction to the technical staff of a high volume, multiple-user research and diagnostic histology laboratory. This position also oversees the laboratory budget, histological procedures and development and implementation of relevant quality assurance and quality control practices. Duties also include performing routine and special histological procedures on animal tissues. Pullman offers a friendly small-town living environment set in the Palouse country having numerous outdoor and recreational activities. QUALIFICATIONS: (Required) Certification as an ASCP Certified Histotechnician or Histotechnologist: experience as a working histotechnician or histotechnologist in high volume setting: supervisory experience in a multiple-employee laboratory setting; good computer skills; ability to organize and prioritize work assignments; ability to work a flexible schedule, including occasional weekends/evenings; strong interpersonal skills with the ability to communicate effectively in writing and orally; ability to anticipate and manage personnel issues, questions and conflicts effectively; able to tolerate legally permissible exposure to chemicals and hazardous material. (Highly Desired) Experience in standard operating procedure (SOP) development and implementation for laboratory quality assurance and quality control practices; experience trimming gross animal tissues for histologic evaluation; familiarity with accredited laboratory procedures, including working in biosafety level 2 facilities and the coordination and implementation of laboratory health and environmental safety programs. SALARY: Salary range of $50,000-55,000/ year commensurate with experience and education. APPLICATION: Send a letter describing your experience and qualifications for this position, a current, detailed r?sum? and contact information for three professional references (include name, addresses, telephone numbers and email addresses) to: Ms. Jeanne Burritt College of Veterinary Medicine Washington State University PO Box 647010 Pullman, WA 99164-7010 Phone: 509-335-3383, Fax: 335-6128 or jburritt@wsu.edu A complete position description may be requested via mail, fax or email. To ensure consideration applications should be received by October 31, 2005. WASHINGTON STATE UNIVERSITY IS AN EQUAL OPPORTUNITY/AFFIRMATIVE ACTION EDUCATOR AND EMPLOYER. Members of ethnic minorities, women, Vietnam-era disabled veterans, persons of disability, and/or persons age 40 and over are encouraged to apply. WSU employs only U.S. citizens and lawfully authorized non-U.S. citizens. All new employees must show employment eligibility verification as required by the U.S. Citizenship and Immigration Services. From David.Edmondson <@t> christie-tr.nwest.nhs.uk Fri Oct 28 11:23:52 2005 From: David.Edmondson <@t> christie-tr.nwest.nhs.uk (Edmondson David (RBV) NHS Christie Tr) Date: Fri Oct 28 11:25:35 2005 Subject: [Histonet] RNase contamination during in-situ Message-ID: <3C83687E8F6AE04792E361ABE2D385B8418056@cht-mail2-2k.xchristie.nhs.uk> 30% Hydrogen peroxide diluted 1 in 10, for ten minutes and rinse glassware with Methanol,,, was what we were advised to use and the slides appear to be not a problem. Do folks agree?? David Manchester, UK The reagents in the kit should not be a problem. The wash/diluent water could be double distilled but the high grade supply for the Bayer analyser in Biochem seems fine -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Barnhart, Tammy Sent: 28 October 2005 16:31 To: Histonet (E-mail) Subject: [Histonet] RNase contamination during in-situ I need some help...we are going to try using the Dako PNA ISH kit for Kappa/Lambda and are concerned about the RNase contamination issue. Can anyone give me an idea as to what product to use to take care of this on surfaces, glassware and reagents? Any and all suggestions would be appreciated. Tammy Barnhart, BS, HTL(ASCP) Anatomic Pathology Supervisor St. Alexius Medical Center Bismarck, ND Confidentiality Notice:This e-mail message is for sole use of intended recipient(s) and may contain confidential and privileged information. Any unauthorized review, use, disclosure, distribution, or copying is prohibited. If you are not the intended recipient, please contact the sender by replying to this e-mail and destroy/delete all copies of this e-mail message. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ****************************************************************************************************************** This e-mail and any files transmitted with it are confidential and solely for the use of the intended recipient. If you have received this e-mail in error you should not disseminate, distribute or copy it. Please notify the sender immediately and delete this e-mail from your system. ******************************************************************************************************************* From awatanabe <@t> tgen.org Fri Oct 28 11:48:09 2005 From: awatanabe <@t> tgen.org (Aprill Watanabe) Date: Fri Oct 28 11:48:25 2005 Subject: [Histonet] Re: Decloaking chamber In-Reply-To: <20051028164214.D3FF420000A4@mr1.tgen.org> Message-ID: Wen, We use it all the time at 15-20 min. The problem is you have to decrease the temperature to go that long. You should be able to press the "display" button until you see SP1 lit up then press the arrow keys for the appropriate time and/or temperature. You can do this for any of the settings to optimize for the tissue you are using. Aprill Watanabe Histotechnologist Tissue Microarray Center Translational Genomics Research Institute (TGen) 602-343-8822 awatanabe@tgen.org On 10/28/05 9:42 AM, "histonet-request@lists.utsouthwestern.edu" wrote: > Send Histonet mailing list submissions to > histonet@lists.utsouthwestern.edu > > To subscribe or unsubscribe via the World Wide Web, visit > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > or, via email, send a message with subject or body 'help' to > histonet-request@lists.utsouthwestern.edu > > You can reach the person managing the list at > histonet-owner@lists.utsouthwestern.edu > > When replying, please edit your Subject line so it is more specific > than "Re: Contents of Histonet digest..." > > > Today's Topics: > > 1. Decloaking Chamber (high pressure cooker) (wen eng) > 2. Help!! We have a problem with a Mounting Media (Gayle Callis) > 3. mouse heart matrix (Kim Merriam) > 4. Re: Decloaking Chamber (high pressure cooker) (Jackie M O'Connor) > 5. RE: Decloaking Chamber (high pressure cooker) (Sebree Linda A.) > 6. Re: Decloaking Chamber (high pressure cooker) (Jan Shivers) > 7. Re: Red HRP chromagen/substrate (Johnson, Teri) > 8. Re: Cholesterol staining suggestions (John A. Kiernan) > 9. Re: mouse heart matrix (Rocan) > 10. RE: Decloaking Chamber (high pressure cooker) > (Favara, Cynthia (NIH/NIAID)) > 11. Mouse Tissue Processing Problems Help (Donna Harclerode) > 12. collagen embeding (Niloufar Fozouni) > 13. FW: [Histonet] mouse heart matrix (Bugelski, Peter [CNTUS]) > 14. Re: questions about rhodanine-copper stain (Mildred Fail) > 15. Thanks for your help! (wen eng) > 16. RE: Help!! We have a problem with a Mounting Media (Gayle Callis) > 17. Solvent Recyclers (Rod Coombe) > 18. VEGF receptor antibodies (Andrea T. Hooper) > 19. Moh's information (Jodiputnam@aol.com) > 20. Moh's information (Stephen Peters M.D.) > 21. Re: Moh's information (John A. Kiernan) > 22. Re: collagen embeding (John A. Kiernan) > 23. IHC Punctate staining in colon epithelium, nonspecific? > (Connolly, Brett M) > 24. RE: Moh's information (Ingles Claire) > 25. RNase contamination during in-situ (Barnhart, Tammy) > 26. New Position (Bailey, Gisela) > 27. RE: RNase contamination during in-situ > (Edmondson David (RBV) NHS Christie Tr) > > > ---------------------------------------------------------------------- > > Message: 1 > Date: Thu, 27 Oct 2005 10:27:49 -0700 (PDT) > From: wen eng > Subject: [Histonet] Decloaking Chamber (high pressure cooker) > To: histonet > Message-ID: <20051027172749.97588.qmail@web53407.mail.yahoo.com> > Content-Type: text/plain; charset=iso-8859-1 > > Hello, > > Does anybody use Decloaking Chamber from Biocare Medical? How should I set the > SP1 temperature and time if I need 20 minutes treatment? The factory setting > is 125 degree 30 seconds and the tech support said 20 minutes is too long. She > suggested me to keep 30 seconds. I still want the suggestion from here. How do > you set your program? > > Thanks! > Wen > > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > > ------------------------------ > > Message: 2 > Date: Thu, 27 Oct 2005 11:30:02 -0600 > From: Gayle Callis > Subject: [Histonet] Help!! We have a problem with a Mounting Media > To: Histonet@lists.utsouthwestern.edu > Message-ID: > <6.0.0.22.1.20051027111608.01b0de68@gemini.msu.montana.edu> > Content-Type: text/plain; charset="us-ascii"; format=flowed > > Is anyone experiencing this problem with Protocol Mounting Medium T, > toluene solvent based from Fisher. > > They take 50 mls and put it into a bottle with a secure, air tight, solvent > resistant lid in order to not dip into stock bottle all the time. > If they use the mounting media absolutely fresh after aliquoting, the > mounting media drys nicely, everything is fine. They are coverslipping > cytospin stained blood cells, Diff Quik stained, and dry without any > immersion oil used before coverslipping. > > If they let the Protocol media aliquot, even though in an air tight, new > bottle, sit around, they notice a layer or some type of separation within > top layer of media. When they use this now separated aliquot, stirred, > and then try to dry slide, the coverslip still slips and slides around - > failure of drying!! Very messy! > > Something is happening, either evaporation of toluene to change ratio of > solvent to acrylic resin, maybe something else i.e faulty > manufacturing/preparation of this media? This is a total mystery to me as > I have done this type of aliquoting with other mounting medias for years > and NEVER had a problem except to add solvent to restore media - thick to > thinner. Nor I have never experienced layering/separation phenomena with > my other mounting media (Richard Allan brand or Permount). After > coverslipping, the latter mounting media set up and dry nicely with no > slip/sliding away!!! > > > > Gayle Callis > Research Histopathology Supervisor > Veterinary Molecular Biology > Montana State University - Bozeman > PO Box 173610 > Bozeman MT 59717-3610 > 406 994-6367 > 406 994-4303 (FAX) > > > > > > ------------------------------ > > Message: 3 > Date: Thu, 27 Oct 2005 10:31:19 -0700 (PDT) > From: Kim Merriam > Subject: [Histonet] mouse heart matrix > To: Histonet > Message-ID: <20051027173119.75498.qmail@web50310.mail.yahoo.com> > Content-Type: text/plain; charset=iso-8859-1 > > Hello all, > > I was wondering if anyone knew of a vendor that sells a tissue matrix that > will allow me to take thin cross-sections of mouse heart (something similar to > the brain matrices sold by Ted Pella. > > Regards, > Kim > > > Kim Merriam > Novartis > Cambridge, MA > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > > ------------------------------ > > Message: 4 > Date: Thu, 27 Oct 2005 12:34:37 -0500 > From: "Jackie M O'Connor" > Subject: Re: [Histonet] Decloaking Chamber (high pressure cooker) > To: wen eng > Cc: histonet > Message-ID: > Content-Type: text/plain; charset="us-ascii" > > I'm using the factory settings for virtually all antibodies - so far, I > haven't found the need to change the settings for anything. I did uncover > a problem, however, with the computer chip in my old Decloaker - the > temperature and pressure were not reaching the set point - but I didn't > notice until I sat down to actually watch the machine. The electronic > temperature display was bouncing all over the place. I had to get a new > machine. > Jackie O' > > > > > > wen eng > Sent by: histonet-bounces@lists.utsouthwestern.edu > 10/27/2005 12:27 PM > > > To: histonet > cc: (bcc: Jackie M O'Connor/LAKE/GPRD/ABBOTT) > Subject: [Histonet] Decloaking Chamber (high pressure cooker) > > > Hello, > > Does anybody use Decloaking Chamber from Biocare Medical? How should I set > the SP1 temperature and time if I need 20 minutes treatment? The factory > setting is 125 degree 30 seconds and the tech support said 20 minutes is > too long. She suggested me to keep 30 seconds. I still want the suggestion > from here. How do you set your program? > > Thanks! > Wen > > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > > > ------------------------------ > > Message: 5 > Date: Thu, 27 Oct 2005 12:43:23 -0500 > From: "Sebree Linda A." > Subject: RE: [Histonet] Decloaking Chamber (high pressure cooker) > To: "wen eng" , "histonet" > > Message-ID: > > Content-Type: text/plain; charset="us-ascii" > > We translated 20 minutes HIER in a water bath to around 2-3 minutes in > the Decloaking Chamber. Hope this helps. > > Linda A. Sebree > University of Wisconsin Hospital & Clinics > IHC/ISH Clinical & Research Laboratory > DM223-VA > 600 Highland Ave. > Madison, WI 53792 > (608)265-6596 > FAX: (608)262-7174 > > > > -----Original Message----- > From: histonet-bounces@lists.utsouthwestern.edu > [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of wen eng > Sent: Thursday, October 27, 2005 12:28 PM > To: histonet > Subject: [Histonet] Decloaking Chamber (high pressure cooker) > > > Hello, > > Does anybody use Decloaking Chamber from Biocare Medical? How should I > set the SP1 temperature and time if I need 20 minutes treatment? The > factory setting is 125 degree 30 seconds and the tech support said 20 > minutes is too long. She suggested me to keep 30 seconds. I still want > the suggestion from here. How do you set your program? > > Thanks! > Wen > > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > > ------------------------------ > > Message: 6 > Date: Thu, 27 Oct 2005 13:01:35 -0500 > From: "Jan Shivers" > Subject: Re: [Histonet] Decloaking Chamber (high pressure cooker) > To: "wen eng" > Cc: histonet > Message-ID: <005901c5db20$78b82e60$41065486@auxs.umn.edu> > Content-Type: text/plain; charset="iso-8859-1" > > I use a Decloaking Chamber for Chronic Wasting Disease slides, and yes, 20 > minutes is the correct setting for that particular test, with a 25 minute > cooldown. Most other IHC tests don't require a lengthy setting on the > Decloakers. > > I'd recommend contacting Biocare to do this for you. They came and > installed the settings on my chambers. > > Jan Shivers > U of MN Vet Diag Lab > > ----- Original Message ----- > From: "wen eng" > To: "histonet" > Sent: Thursday, October 27, 2005 12:27 PM > Subject: [Histonet] Decloaking Chamber (high pressure cooker) > > >> Hello, >> >> Does anybody use Decloaking Chamber from Biocare Medical? How should I set > the SP1 temperature and time if I need 20 minutes treatment? The factory > setting is 125 degree 30 seconds and the tech support said 20 minutes is too > long. She suggested me to keep 30 seconds. I still want the suggestion from > here. How do you set your program? >> >> Thanks! >> Wen >> >> >> --------------------------------- >> Yahoo! FareChase - Search multiple travel sites in one click. >> _______________________________________________ >> Histonet mailing list >> Histonet@lists.utsouthwestern.edu >> http://lists.utsouthwestern.edu/mailman/listinfo/histonet >> > > > > > ------------------------------ > > Message: 7 > Date: Thu, 27 Oct 2005 14:24:14 -0500 > From: "Johnson, Teri" > Subject: [Histonet] Re: Red HRP chromagen/substrate > To: > Message-ID: > > > Content-Type: text/plain; charset="us-ascii" > > Andrea, > > It depends on what you consider "best". As Gayle pointed out, for some > it's endpoint color. For others, it's permanence. AEC needs to be > mounted in aqueous mountant because it's not stable in organic solvents > (even the polyvinyl alcohol in some of the aqueous mountants can cause > fading!). The way to get around this is to either use a permanent > chromogen (Vector Nova Red), or you can use Crystal mount to cover the > AEC stained tissue sample and allow to dry. We always coverslip it with > permanent mountant for better visualization under the microscope. > > Teri Johnson > Managing Director Histology Facility > Stowers Institute for Medical Research > 1000 E. 50th St. > Kansas City, MO 64133 > > > > ------------------------------ > > Message: 8 > Date: Thu, 27 Oct 2005 15:36:27 -0400 > From: "John A. Kiernan" > Subject: Re: [Histonet] Cholesterol staining suggestions > To: Gayle Callis > Cc: Histonet@lists.utsouthwestern.edu > Message-ID: <43612C3B.BD853396@uwo.ca> > Content-Type: text/plain; charset=us-ascii > > The only specific method for cholesterol that > doesn't involve a nasty acid is the cholesterol > oxidase-DAB method (Emeis et al 1977 Histochem. J. > 9:197-204). It can also detect cholesterol esters > if the sections are first treated with cholesterol > esterase. It's rather more than a "stain" but should > be OK for research. It can also be used at the EM > level (eg Pelletier & Vitale 1994 J Histochem Cytochem > 42:1539-1554.) > > Regarding the traditional stains for cholesterol: > According to Adams, Pearse and Bayliss High, the > perchloric acid-naphthoquinone method is the most > specific and sensitive one for cholesterol and its > esters. I've tried it once or twice; it works but > it's rather rough on the sections. The blue colour > fades after a few hours. An older, simpler method > is that of Schultz (here summarized from Bayliss High's > 1984 "Lipid Histochemistry" p.40-41). I've not tried > it myself. > > Frozen sections, after formal-calcium fixation, > air-dried onto slides. > 1. Oxidize sections in 2.5% iron alum in 0.2M acetate > buffer, pH 3 for 4 hours at 37C. > 2. Wash, 3 X 20 min in the acetate buffer. > 3. Rinse in distilled water. > 4. Treat with 5% formalin in tap water > (ie 2% formaldehyde), 10 min. > 5. Drain and allow to dry. > 6. Apply a drop or two of Schulz reagent (see below), > and then a coverslip. Examine with microscope. > Result. Cholesterol blue-green, fading quickly. > Schulz reagent: Add 1ml glacial acetic acid dropwise to > 1 ml conc. sulphuric acid, with agitation. It becomes hot > and syrupy; let it cool. > > Cholesterol and its esters are birefringent with crossed > polars. This property was discovered by B. Doinikow (1913) > Deutsche Z. Nervenheilk. 46:20-42) and it has been exploited > to trace degenerating myelinated axons in frozen sections of > human brain (Miklossy & Van der Loos 1987 Brain Res. > 426:377-380). > I tried to reproduce their results in the 1990s but the > sections contained great quantities of birefringent dirt > (completely invisible with ordinary illumination). The dirt > didn't consist of tiny needle-like crystals like the ones > described by Doinikow and shown in the colour photos in > Miklossy's paper. My material had been stored for a long > time > in formalin before cutting, so perhaps lipids everywhere > had changed into some sort of birefringent junk. From info <@t> instrumedics.com Fri Oct 28 12:15:25 2005 From: info <@t> instrumedics.com (Instrumedics) Date: Fri Oct 28 12:15:52 2005 Subject: [Histonet] Mohs information Message-ID: <018301c5dbe3$399b37e0$6401a8c0@INSTRUMEDICS22> Many Mohs laboratories use the Gentle Jane for snap freezing the skin specimen. The tissue is flat with the margins in the same plane and at the surface of the blockface. Snap-freezing with a heat extractor chilled in liquid nitrogen provides a very cold temperature with good thermal exchange. The ice crystal growth is minimized and the morphology is preserved . Please visit www.instrumedics.com and click on "snap freezing" . There is a step-by-step demonstration of the Gentle Jane freezing process. Bernice From anh2006 <@t> med.cornell.edu Fri Oct 28 12:36:02 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Fri Oct 28 12:31:12 2005 Subject: [Histonet] Paraffins for mouse tissue Message-ID: What are the best paraffins to use for infiltrating and embedding mouse tissues. I currently use one for infiltrating and another for embedding but I would love to see what others are using and recommend to be the best. Thanks, Andrea -- From ASuarez <@t> mrl.ubc.ca Fri Oct 28 12:51:14 2005 From: ASuarez <@t> mrl.ubc.ca (Agripina Suarez) Date: Fri Oct 28 12:51:45 2005 Subject: [Histonet] RNase contamination during in-situ In-Reply-To: <1779904B5E82D511914C00D0B793339205BFDA6F@exchangent> References: <1779904B5E82D511914C00D0B793339205BFDA6F@exchangent> Message-ID: <436202A2020000E2000007CD@mail.mrl.ubc.ca> I use DEPC water for all solutions. All glassware are baked at 200oC for at least 200oC. I also use RNaseZap (Ambion) followed by DEPC water rinse for small items that are not RNase-free. I also purchase RNase-free plasticware. Agripina C. Suarez McDonald Research Laboratories iCAPTURE Centre, St Paul's Hospital 1081 Burrard St, Vancouver B.C., Canada V6Z 1Y6 Tel no. 604 682 2344 local 62562 Fax no. 604 806 8351 >>> "Barnhart, Tammy" 10/28/05 8:31 AM >>> I need some help...we are going to try using the Dako PNA ISH kit for Kappa/Lambda and are concerned about the RNase contamination issue. Can anyone give me an idea as to what product to use to take care of this on surfaces, glassware and reagents? Any and all suggestions would be appreciated. Tammy Barnhart, BS, HTL(ASCP) Anatomic Pathology Supervisor St. Alexius Medical Center Bismarck, ND Confidentiality Notice:This e-mail message is for sole use of intended recipient(s) and may contain confidential and privileged information. Any unauthorized review, use, disclosure, distribution, or copying is prohibited. If you are not the intended recipient, please contact the sender by replying to this e-mail and destroy/delete all copies of this e-mail message. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From gcallis <@t> montana.edu Fri Oct 28 12:54:09 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Fri Oct 28 12:54:21 2005 Subject: [Histonet] Paraffins for mouse tissue In-Reply-To: References: Message-ID: <6.0.0.22.1.20051028114652.01b4fd00@gemini.msu.montana.edu> No one is superior, in my opinion, having tried just about everything on the market over the years. We have used Surgipath's infiltration media and their embedding media combination/system with great success. We have gone back to Fishers Tissue Prep 2, consensus of opinion for mouse and hamster brains, decalcified mouse bones, and all other murine tissues, we like it best for both infiltration and embedding. We are using up Tissue Prep from Fisher and find it bit softer but adequate for infiltration and embedding. In general, once we chose a paraffin, we use it for both infiltration and embedding with the exception of Surgipaths tidy combo system. I think the key is how mouse tissue is processed before the paraffin infiltration/embedding, if not done optimally, no paraffin has bailed us out of trouble nor gives us an opinion on what is best. At 11:36 AM 10/28/2005, you wrote: >What are the best paraffins to use for infiltrating and embedding mouse >tissues. I currently use one for infiltrating and another for embedding >but I would love to see what others are using and recommend to be the best. > >Thanks, >Andrea >-- > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From anh2006 <@t> med.cornell.edu Fri Oct 28 12:59:18 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Fri Oct 28 12:54:29 2005 Subject: [Histonet] Hand processing mouse tissues - schedule needed Message-ID: I would love to hear your protocols for processing mouse tissue by hand. We are thinking of trying new protocols. Thanks, Andrea -- From ryaskovich <@t> dir.nidcr.nih.gov Fri Oct 28 13:10:09 2005 From: ryaskovich <@t> dir.nidcr.nih.gov (Yaskovich, Ruth A (NIH/NIDCR)) Date: Fri Oct 28 13:10:19 2005 Subject: [Histonet] Hand processing mouse tissues - schedule needed Message-ID: I have processed all kinds of rodent tissue by hand. Sometimes I haven't even remembered the times I had it in each alcohol or xylene or paraffin. It's always come out just fine no cutting problems. I think as long as it's fixed well it doesn't matter. Ruth Yaskovich National Institutes of Health National Institute of Dental and Crainiofacial Research Pain and Neurosensory Mechanisms Branch Drug Discovery > ---------- > From: Andrea T. Hooper > Sent: Friday, October 28, 2005 12:59 PM > To: Histonet > Subject: [Histonet] Hand processing mouse tissues - schedule needed > > I would love to hear your protocols for processing mouse tissue by > hand. We are thinking of trying new protocols. > > Thanks, > Andrea > -- > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > From jcox90 <@t> yahoo.com Fri Oct 28 13:46:25 2005 From: jcox90 <@t> yahoo.com (Jill Cox) Date: Fri Oct 28 13:46:34 2005 Subject: [Histonet] Di water for Pathology lab Message-ID: <20051028184625.70437.qmail@web52110.mail.yahoo.com> Hello Histonetters, We are purchasing a deionized water system for our new pathology lab. Does anyone have a good company they can recommend? Would love any comments as well, Thanks in advance, Jill Jill Cox HT (ASCP) From ROrr <@t> enh.org Fri Oct 28 13:53:04 2005 From: ROrr <@t> enh.org (Orr, Rebecca) Date: Fri Oct 28 13:53:14 2005 Subject: [Histonet] Decloaker Message-ID: Wen, I have used the Biocare Decloaker for several years. In my experience, I would agree 20 minutes is really too long, at such a high temperature. You will most likely get staining, but you will most definitely loose tissue as well as morphology. I know of labs like Jan Shivers' at U of MN that use the decloaker to test prions and that has an extended time at the 120'-125'C. So there are applications where this may be beneficial to you... But clinically speaking, if you have to work that hard, something's wrong somewhere else! I understand research and non hospital settings have different needs and unique antibodies...but if I would have to use, in my opinion, such extreme parameters, I would try a different ph solution, or look at the enzymes to digest or even a stronger dilution. Then there's always fixation issues... >From what I get from your email is that you asked for directions to extend the time at the high temperature. I'm sure Biocare Tech support instructed you how to make the change with the recommendation that you shouldn't really need that, for HEIR. You've gotten a bunch of our opinions, as to why you shouldn't need that high a temp, but regardless, it's your lab and your test and your responsibility...so have at it! And let us know your results! As to how to set the program, I see others have shared. Good luck, I'd be very interested to hear of your results. Becky Orr CLA,HT(ASCP) IHC Lead Evanston Northwestern Healthcare 847-570-2771 > Hello, > > Does anybody use Decloaking Chamber from Biocare Medical? How should I set the > SP1 temperature and time if I need 20 minutes treatment? The factory setting > is 125 degree 30 seconds and the tech support said 20 minutes is too long. She > suggested me to keep 30 seconds. I still want the suggestion from here. How do > you set your program? > > Thanks! > Wen > > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > > ------------------------------ > > Message: 2 > Date: Thu, 27 Oct 2005 11:30:02 -0600 > From: Gayle Callis > Subject: [Histonet] Help!! We have a problem with a Mounting Media > To: Histonet@lists.utsouthwestern.edu > Message-ID: > <6.0.0.22.1.20051027111608.01b0de68@gemini.msu.montana.edu> > Content-Type: text/plain; charset="us-ascii"; format=flowed > > Is anyone experiencing this problem with Protocol Mounting Medium T, > toluene solvent based from Fisher. > > They take 50 mls and put it into a bottle with a secure, air tight, solvent > resistant lid in order to not dip into stock bottle all the time. > If they use the mounting media absolutely fresh after aliquoting, the > mounting media drys nicely, everything is fine. They are coverslipping > cytospin stained blood cells, Diff Quik stained, and dry without any > immersion oil used before coverslipping. > > If they let the Protocol media aliquot, even though in an air tight, new > bottle, sit around, they notice a layer or some type of separation within > top layer of media. When they use this now separated aliquot, stirred, > and then try to dry slide, the coverslip still slips and slides around - > failure of drying!! Very messy! > > Something is happening, either evaporation of toluene to change ratio of > solvent to acrylic resin, maybe something else i.e faulty > manufacturing/preparation of this media? This is a total mystery to me as > I have done this type of aliquoting with other mounting medias for years > and NEVER had a problem except to add solvent to restore media - thick to > thinner. Nor I have never experienced layering/separation phenomena with > my other mounting media (Richard Allan brand or Permount). After > coverslipping, the latter mounting media set up and dry nicely with no > slip/sliding away!!! > > > > Gayle Callis > Research Histopathology Supervisor > Veterinary Molecular Biology > Montana State University - Bozeman > PO Box 173610 > Bozeman MT 59717-3610 > 406 994-6367 > 406 994-4303 (FAX) > > > > > > ------------------------------ > > Message: 3 > Date: Thu, 27 Oct 2005 10:31:19 -0700 (PDT) > From: Kim Merriam > Subject: [Histonet] mouse heart matrix > To: Histonet > Message-ID: <20051027173119.75498.qmail@web50310.mail.yahoo.com> > Content-Type: text/plain; charset=iso-8859-1 > > Hello all, > > I was wondering if anyone knew of a vendor that sells a tissue matrix that > will allow me to take thin cross-sections of mouse heart (something similar to > the brain matrices sold by Ted Pella. > > Regards, > Kim > > > Kim Merriam > Novartis > Cambridge, MA > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > > ------------------------------ > > Message: 4 > Date: Thu, 27 Oct 2005 12:34:37 -0500 > From: "Jackie M O'Connor" > Subject: Re: [Histonet] Decloaking Chamber (high pressure cooker) > To: wen eng > Cc: histonet > Message-ID: > Content-Type: text/plain; charset="us-ascii" > > I'm using the factory settings for virtually all antibodies - so far, I > haven't found the need to change the settings for anything. I did uncover > a problem, however, with the computer chip in my old Decloaker - the > temperature and pressure were not reaching the set point - but I didn't > notice until I sat down to actually watch the machine. The electronic > temperature display was bouncing all over the place. I had to get a new > machine. > Jackie O' > > > > > > wen eng > Sent by: histonet-bounces@lists.utsouthwestern.edu > 10/27/2005 12:27 PM > > > To: histonet > cc: (bcc: Jackie M O'Connor/LAKE/GPRD/ABBOTT) > Subject: [Histonet] Decloaking Chamber (high pressure cooker) > > > Hello, > > Does anybody use Decloaking Chamber from Biocare Medical? How should I set > the SP1 temperature and time if I need 20 minutes treatment? The factory > setting is 125 degree 30 seconds and the tech support said 20 minutes is > too long. She suggested me to keep 30 seconds. I still want the suggestion > from here. How do you set your program? > > Thanks! > Wen > > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > > > ------------------------------ > > Message: 5 > Date: Thu, 27 Oct 2005 12:43:23 -0500 > From: "Sebree Linda A." > Subject: RE: [Histonet] Decloaking Chamber (high pressure cooker) > To: "wen eng" , "histonet" > > Message-ID: > > Content-Type: text/plain; charset="us-ascii" > > We translated 20 minutes HIER in a water bath to around 2-3 minutes in > the Decloaking Chamber. Hope this helps. > > Linda A. Sebree > University of Wisconsin Hospital & Clinics > IHC/ISH Clinical & Research Laboratory > DM223-VA > 600 Highland Ave. > Madison, WI 53792 > (608)265-6596 > FAX: (608)262-7174 > > > > -----Original Message----- > From: histonet-bounces@lists.utsouthwestern.edu > [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of wen eng > Sent: Thursday, October 27, 2005 12:28 PM > To: histonet > Subject: [Histonet] Decloaking Chamber (high pressure cooker) > > > Hello, > > Does anybody use Decloaking Chamber from Biocare Medical? How should I > set the SP1 temperature and time if I need 20 minutes treatment? The > factory setting is 125 degree 30 seconds and the tech support said 20 > minutes is too long. She suggested me to keep 30 seconds. I still want > the suggestion from here. How do you set your program? > > Thanks! > Wen > > > --------------------------------- > Yahoo! FareChase - Search multiple travel sites in one click. > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > > ------------------------------ > > Message: 6 > Date: Thu, 27 Oct 2005 13:01:35 -0500 > From: "Jan Shivers" > Subject: Re: [Histonet] Decloaking Chamber (high pressure cooker) > To: "wen eng" > Cc: histonet > Message-ID: <005901c5db20$78b82e60$41065486@auxs.umn.edu> > Content-Type: text/plain; charset="iso-8859-1" > > I use a Decloaking Chamber for Chronic Wasting Disease slides, and yes, 20 > minutes is the correct setting for that particular test, with a 25 minute > cooldown. Most other IHC tests don't require a lengthy setting on the > Decloakers. > > I'd recommend contacting Biocare to do this for you. They came and > installed the settings on my chambers. > > Jan Shivers > U of MN Vet Diag Lab > > ----- Original Message ----- > From: "wen eng" > To: "histonet" > Sent: Thursday, October 27, 2005 12:27 PM > Subject: [Histonet] Decloaking Chamber (high pressure cooker) > > >> Hello, >> >> Does anybody use Decloaking Chamber from Biocare Medical? How should I set > the SP1 temperature and time if I need 20 minutes treatment? The factory > setting is 125 degree 30 seconds and the tech support said 20 minutes is too > long. She suggested me to keep 30 seconds. I still want the suggestion from > here. How do you set your program? >> >> Thanks! >> Wen >> >> >> --------------------------------- >> Yahoo! FareChase - Search multiple travel sites in one click. >> _______________________________________________ >> Histonet mailing list >> Histonet@lists.utsouthwestern.edu >> http://lists.utsouthwestern.edu/mailman/listinfo/histonet >> > > > > > ------------------------------ > > Message: 7 > Date: Thu, 27 Oct 2005 14:24:14 -0500 > From: "Johnson, Teri" > Subject: [Histonet] Re: Red HRP chromagen/substrate > To: > Message-ID: > > > Content-Type: text/plain; charset="us-ascii" > > Andrea, > > It depends on what you consider "best". As Gayle pointed out, for some > it's endpoint color. For others, it's permanence. AEC needs to be > mounted in aqueous mountant because it's not stable in organic solvents > (even the polyvinyl alcohol in some of the aqueous mountants can cause > fading!). The way to get around this is to either use a permanent > chromogen (Vector Nova Red), or you can use Crystal mount to cover the > AEC stained tissue sample and allow to dry. We always coverslip it with > permanent mountant for better visualization under the microscope. > > Teri Johnson > Managing Director Histology Facility > Stowers Institute for Medical Research > 1000 E. 50th St. > Kansas City, MO 64133 > > > > ------------------------------ > > Message: 8 > Date: Thu, 27 Oct 2005 15:36:27 -0400 > From: "John A. Kiernan" > Subject: Re: [Histonet] Cholesterol staining suggestions > To: Gayle Callis > Cc: Histonet@lists.utsouthwestern.edu > Message-ID: <43612C3B.BD853396@uwo.ca> > Content-Type: text/plain; charset=us-ascii > > The only specific method for cholesterol that > doesn't involve a nasty acid is the cholesterol > oxidase-DAB method (Emeis et al 1977 Histochem. J. > 9:197-204). It can also detect cholesterol esters > if the sections are first treated with cholesterol > esterase. It's rather more than a "stain" but should > be OK for research. It can also be used at the EM > level (eg Pelletier & Vitale 1994 J Histochem Cytochem > 42:1539-1554.) > > Regarding the traditional stains for cholesterol: > According to Adams, Pearse and Bayliss High, the > perchloric acid-naphthoquinone method is the most > specific and sensitive one for cholesterol and its > esters. I've tried it once or twice; it works but > it's rather rough on the sections. The blue colour > fades after a few hours. An older, simpler method > is that of Schultz (here summarized from Bayliss High's > 1984 "Lipid Histochemistry" p.40-41). I've not tried > it myself. > > Frozen sections, after formal-calcium fixation, > air-dried onto slides. > 1. Oxidize sections in 2.5% iron alum in 0.2M acetate > buffer, pH 3 for 4 hours at 37C. > 2. Wash, 3 X 20 min in the acetate buffer. > 3. Rinse in distilled water. > 4. Treat with 5% formalin in tap water > (ie 2% formaldehyde), 10 min. > 5. Drain and allow to dry. > 6. Apply a drop or two of Schulz reagent (see below), > and then a coverslip. Examine with microscope. > Result. Cholesterol blue-green, fading quickly. > Schulz reagent: Add 1ml glacial acetic acid dropwise to > 1 ml conc. sulphuric acid, with agitation. It becomes hot > and syrupy; let it cool. > > Cholesterol and its esters are birefringent with crossed > polars. This property was discovered by B. Doinikow (1913) > Deutsche Z. Nervenheilk. 46:20-42) and it has been exploited > to trace degenerating myelinated axons in frozen sections of > human brain (Miklossy & Van der Loos 1987 Brain Res. > 426:377-380). > I tried to reproduce their results in the 1990s but the > sections contained great quantities of birefringent dirt > (completely invisible with ordinary illumination). The dirt > didn't consist of tiny needle-like crystals like the ones > described by Doinikow and shown in the colour photos in > Miklossy's paper. My material had been stored for a long > time > in formalin before cutting, so perhaps lipids everywhere > had changed into some sort of birefringent junk. ------------------------------ _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet End of Histonet Digest, Vol 23, Issue 33 **************************************** From Jackie.O'Connor <@t> abbott.com Fri Oct 28 14:41:54 2005 From: Jackie.O'Connor <@t> abbott.com (Jackie M O'Connor) Date: Fri Oct 28 14:42:21 2005 Subject: [Histonet] Chemicon Tissue Microarray Message-ID: Does anyone have one? I have a Beecher manual right now - is the Chemicon Arrayer worth the additional expense? It doesn't look to be much better - I'd appreciate some insight. Jackie O' From quinntl <@t> umkc.edu Fri Oct 28 14:50:51 2005 From: quinntl <@t> umkc.edu (Quinn, Tim L.) Date: Fri Oct 28 14:51:04 2005 Subject: [Histonet] Autofluorescense of rbc's Message-ID: <3563FA9377A5DC41BDDA1E8A6C06BEF803E9C0CD@KC-MAIL6.kc.umkc.edu> Dear listies, I'm sure you've had this inquiry before but I'm somewhat of a newbee. I'm attempting to use fluorescent markers on paraffin sections and the rbc's are autofluoresing. What is the best method for removing the fluorescing components? Thanks. Timothy L. Quinn Senior Lab Techician Missouri University at Kansas City Medical School- Med Lab 2411 Holmes Kansas City, MO 64108 816-235-1904 quinntl@umkc.edu From pruegg <@t> ihctech.net Fri Oct 28 15:08:59 2005 From: pruegg <@t> ihctech.net (Patsy Ruegg) Date: Fri Oct 28 15:09:16 2005 Subject: [Histonet] alcian blue/pas Message-ID: <200510282008.j9SK8v6m023089@chip.viawest.net> Can someone steer me to a good alcian blue/pas stain for frozen sections. We have mouse tissue labeled with Lac Z and someone mentioned that cells making mucous could actually non-specifically stain with the blue Lac Z, so we need to prove the positive stain we have with Lac Z is real and not because the cells are making mucous. Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech, LLC Fitzsimmons BioScience Park 12635 Montview Blvd. Suite 216 Aurora, CO 80010 P-720-859-4060 F-720-859-4110 wk email pruegg@ihctech.net web site www.ihctech.net This email is confidential and intended solely for the use of the Person(s) ('the intended recipient') to whom it was addressed. Any views or opinions presented are solely those of the author. It may contain information that is privileged & confidential within the meaning of applicable law. Accordingly any dissemination, distribution, copying, or other use of this message, or any of its contents, by any person other than the intended recipient may constitute a breach of civil or criminal law and is strictly prohibited. If you are NOT the intended recipient please contact the sender and dispose of this e-mail as soon as possible. From gcallis <@t> montana.edu Fri Oct 28 15:59:18 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Fri Oct 28 15:59:39 2005 Subject: [Histonet] alcian blue/pas In-Reply-To: <200510282008.j9SK8v6m023089@chip.viawest.net> References: <200510282008.j9SK8v6m023089@chip.viawest.net> Message-ID: <6.0.0.22.1.20051028144750.01b668d8@gemini.msu.montana.edu> What tissue are you staining? Mouse intestine? At 02:08 PM 10/28/2005, you wrote: >Can someone steer me to a good alcian blue/pas stain for frozen sections. >We have mouse tissue labeled with Lac Z and someone mentioned that cells >making mucous could actually non-specifically stain with the blue Lac Z, so >we need to prove the positive stain we have with Lac Z is real and not >because the cells are making mucous. >Patsy > >Patsy Ruegg, HT(ASCP)QIHC >IHCtech, LLC >Fitzsimmons BioScience Park >12635 Montview Blvd. Suite 216 >Aurora, CO 80010 >P-720-859-4060 >F-720-859-4110 >wk email pruegg@ihctech.net >web site www.ihctech.net > > >This email is confidential and intended solely for the use of the Person(s) >('the intended recipient') to whom it was addressed. Any views or opinions >presented are solely those of the author. It may contain information that is >privileged & confidential within the meaning of applicable law. Accordingly >any dissemination, distribution, copying, or other use of this message, or >any of its contents, by any person other than the intended recipient may >constitute a breach of civil or criminal law and is strictly prohibited. If >you are NOT the intended recipient please contact the sender and dispose of >this e-mail as soon as possible. > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From ASuarez <@t> mrl.ubc.ca Fri Oct 28 16:13:46 2005 From: ASuarez <@t> mrl.ubc.ca (Agripina Suarez) Date: Fri Oct 28 16:14:15 2005 Subject: [Histonet] RNase contamination during in-situ In-Reply-To: <436202A2020000E2000007CD@mail.mrl.ubc.ca> References: <1779904B5E82D511914C00D0B793339205BFDA6F@exchangent> <436202A2020000E2000007CD@mail.mrl.ubc.ca> Message-ID: <4362321A020000E2000007F2@mail.mrl.ubc.ca> Ooops. Should read "... baked at 200oC for at least 2 hrours." Agripina C. Suarez McDonald Research Laboratories iCAPTURE Centre, St Paul's Hospital 1081 Burrard St, Vancouver B.C., Canada V6Z 1Y6 Tel no. 604 682 2344 local 62562 Fax no. 604 806 8351 >>> "Agripina Suarez" 10/28/05 10:51 AM >>> I use DEPC water for all solutions. All glassware are baked at 200oC for at least 200oC. I also use RNaseZap (Ambion) followed by DEPC water rinse for small items that are not RNase-free. I also purchase RNase-free plasticware. Agripina C. Suarez McDonald Research Laboratories iCAPTURE Centre, St Paul's Hospital 1081 Burrard St, Vancouver B.C., Canada V6Z 1Y6 Tel no. 604 682 2344 local 62562 Fax no. 604 806 8351 >>> "Barnhart, Tammy" 10/28/05 8:31 AM >>> I need some help...we are going to try using the Dako PNA ISH kit for Kappa/Lambda and are concerned about the RNase contamination issue. Can anyone give me an idea as to what product to use to take care of this on surfaces, glassware and reagents? Any and all suggestions would be appreciated. Tammy Barnhart, BS, HTL(ASCP) Anatomic Pathology Supervisor St. Alexius Medical Center Bismarck, ND Confidentiality Notice:This e-mail message is for sole use of intended recipient(s) and may contain confidential and privileged information. Any unauthorized review, use, disclosure, distribution, or copying is prohibited. If you are not the intended recipient, please contact the sender by replying to this e-mail and destroy/delete all copies of this e-mail message. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From deb.vaneyck <@t> phci.org Fri Oct 28 18:13:05 2005 From: deb.vaneyck <@t> phci.org (Van Eyck, Deb) Date: Fri Oct 28 18:13:19 2005 Subject: [Histonet] amyloid A & P Message-ID: Hi -----does anyone out there have a method of staining fat pad aspiration smears for amyloid a and p that consistently works---I have no problem on formalin fixed blocks but the smears don't always survive the retrieval or digestion employed. I usually formalin fix all my cytology specimens and they survive the same processing that formalin fixed blocks do. Thanks for any input. Deb This information is confidential and intended solely for the use of the individual or entity to whom it is addressed. If you have received this email in error please notify the sender or our Customer Support Center at (262) 928-2777. We have scanned this email and its attachments for malicious content. However, the recipient should check this email and any attachments for the presence of viruses. ProHealth Care accepts no liability for any damage caused by any virus transmitted by this email. From tahseen <@t> brain.net.pk Fri Oct 28 15:31:21 2005 From: tahseen <@t> brain.net.pk (Muhammad Tahseen) Date: Fri Oct 28 23:34:08 2005 Subject: [Histonet] Purcase a microwave tissue processor Message-ID: <015d01c5dbfe$9073a000$972bfea9@m7c0y4> We are looking to purchase a microwave tissue processor for surgical specimens. What has been the experience of users in Histoland as to preferred manufacturers? Muhammad Tahseen Histology Supervisor SKMCH & RC Lahore Pakistan. From jkiernan <@t> uwo.ca Sat Oct 29 23:49:09 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Sat Oct 29 23:49:20 2005 Subject: [Histonet] alcian blue/pas References: <200510282008.j9SK8v6m023089@chip.viawest.net> Message-ID: <436450C5.FD58A88E@uwo.ca> Dear Patsy Ruegg, If you're working with frozen sections of a tissue that contains endogenous beta-galactosidase, the usual method for staining the lac-z gene product should not give a false-positive because the pH optima for the animal (lysosomal) and E. coli galactosidases are widely separated. I'm assumong that you're detecting the marker enzyme with the usual indogogenic method, using the substrate commonly called X-gal. (I can't be bothered to type the real name.) That said, the product of the indigogenic X-gal reaction has a rather unpleasing greenish blue colour. Material stained with alcian blue has a turquoise colour - a lovely balance of blue and green. The X-gal product and alcian blue colours are not the same, but they are similar. There are indigogenic beta-galactosides that yield green and red indigoid dyes, but I have found no published reports of their use with tissue sections and microscopes. Please let me know if I've missed something in this field. The insoluble indigoid pigment formed from X-gal is formed more quickly than the ones from other indoxyl beta-galactosides. The rather dirty-looking blue-green product is deposited near the molecules of the galactosidase. "Near" in this context means in the cytoplasm rather than the nucleus, and perhaps at the basal or apical end of an epithelial cell. This is LM, not EM enzyme histochemistry. The X-gal method is "standard" among molecular biologists. You may want to consider alternatives to an alcian blue & PAS counterstain. The bromo-chloro-indigo product of the X-gal reaction is almost as indestructible as copper phthalocyanine (the product of alcian blue staining). There are several alcian blue-PAS and PAS-alcian blue methods (the sequence affects the result, and in a logical way). What is expected of the counterstain to your lac-z reporter gene? If greeny-blue is mandatory for lac-z localization, all the other colours are up for grabs. John Kiernan Anatomy, UWO London, Canada. _____________________________ Patsy Ruegg wrote: > > Can someone steer me to a good alcian blue/pas stain for frozen sections. > We have mouse tissue labeled with Lac Z and someone mentioned that cells > making mucous could actually non-specifically stain with the blue Lac Z, so > we need to prove the positive stain we have with Lac Z is real and not > because the cells are making mucous. > Patsy > > Patsy Ruegg, HT(ASCP)QIHC > IHCtech, LLC > Fitzsimmons BioScience Park > 12635 Montview Blvd. Suite 216 > Aurora, CO 80010 > P-720-859-4060 > F-720-859-4110 > wk email pruegg@ihctech.net > web site www.ihctech.net > > > This email is confidential and intended solely for the use of the Person(s) > ('the intended recipient') to whom it was addressed. Any views or opinions > presented are solely those of the author. It may contain information that is > privileged & confidential within the meaning of applicable law. Accordingly > any dissemination, distribution, copying, or other use of this message, or > any of its contents, by any person other than the intended recipient may > constitute a breach of civil or criminal law and is strictly prohibited. If > you are NOT the intended recipient please contact the sender and dispose of > this e-mail as soon as possible. > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet From jkiernan <@t> uwo.ca Sat Oct 29 23:50:19 2005 From: jkiernan <@t> uwo.ca (John Kiernan) Date: Sat Oct 29 23:50:27 2005 Subject: [Histonet] Hand processing mouse tissues - schedule needed References: Message-ID: <4364510B.63B4C48F@uwo.ca> Well said! "Yaskovich, Ruth A (NIH/NIDCR)" wrote: > > I have processed all kinds of rodent tissue by hand. Sometimes I haven't even remembered the times I had it in each alcohol or xylene or paraffin. It's always come out just fine no cutting problems. I think as long as it's fixed well it doesn't matter. > Ruth Yaskovich > National Institutes of Health > National Institute of Dental and Crainiofacial Research > Pain and Neurosensory Mechanisms Branch > Drug Discovery > > > > I would love to hear your protocols for processing mouse tissue by > > hand. We are thinking of trying new protocols. From pruegg <@t> ihctech.net Sun Oct 30 08:45:06 2005 From: pruegg <@t> ihctech.net (patsy ruegg) Date: Sun Oct 30 08:45:21 2005 Subject: [Histonet] fibrilin Message-ID: <200510301445.j9UEj3vZ093845@pro12.abac.com> Anybody doing IHC with an antibody to fibrilin on mouse tissue? I am using a rab. Poly. Antibody from Santa Cruz, with limited success. Patsy Ruegg, HT(ASCP)QIHC IHCtech 12635 Montview Blvd. #216 Aurora, CO 80010 720-859-4060 fax 720-859-4110 pruegg@ihctech.net www.ihctech.net From MVaughan4 <@t> ucok.edu Sun Oct 30 12:53:38 2005 From: MVaughan4 <@t> ucok.edu (MVaughan4@ucok.edu) Date: Sun Oct 30 12:54:18 2005 Subject: [Histonet] Salivary gland tissue fetal pig problems In-Reply-To: Message-ID: Hello again, I want to get salivary gland tissue (parotid, submandibular, sublingual) for the students. I thought I would try using the glands from the fetal pigs (sent from Carolina Bio) we use for student dissection. I post-fixed them briefly (~30 minutes) in 10%NBF, followed by dehydration/infiltration. After sectioning, the connective tissue is mostly degenerated and the gland tissue doesn't look that good either. The good archival gland tissue we have is yellowish-colored, maybe it is fixed in something different than NBF. Any suggestions? Thanks. Mel Melville B. Vaughan, Ph. D. Assistant Professor Department of Biology University of Central Oklahoma Edmond, OK 73034 From anh2006 <@t> med.cornell.edu Sun Oct 30 14:29:35 2005 From: anh2006 <@t> med.cornell.edu (Andrea T. Hooper) Date: Sun Oct 30 14:24:50 2005 Subject: [Histonet] alcian blue/pas In-Reply-To: <200510282008.j9SK8v6m023089@chip.viawest.net> References: <200510282008.j9SK8v6m023089@chip.viawest.net> Message-ID: Dear Patsy, The best control for BGal staining is the same tissue type from a wild-type mouse littermate (same strain) that does not contain the gene for LacZ. I assume you are working with transgenic mice here? If not, ignore this comment. Incidentally, what tissue are you staining? Good luck, Andrea >Can someone steer me to a good alcian blue/pas stain for frozen sections. >We have mouse tissue labeled with Lac Z and someone mentioned that cells >making mucous could actually non-specifically stain with the blue Lac Z, so >we need to prove the positive stain we have with Lac Z is real and not >because the cells are making mucous. >Patsy > -- From ASelf <@t> gmhsc.com Mon Oct 31 05:53:26 2005 From: ASelf <@t> gmhsc.com (Amy Self) Date: Mon Oct 31 05:53:42 2005 Subject: [Histonet] Microtome Message-ID: <39836CD6DB61654E8F95A35898C92186D70FFD@exchange.gmhpost.com> Happy Halloween Netters.... Does anyone have a used microtome that they would like to get rid of...... Thanks, amy NOTE: The information contained in this message may be privileged, confidential and protected from disclosure. If the reader of this message is not the intended recipient, or an employee or agent responsible for delivering this message to the intended recipient, you are hereby notified that any dissemination, distribution or copying of this communication is strictly prohibited. If you have received this communication in error, please notify us immediately by replying to this message and deleting it from your computer. Thank you. From mdean <@t> uci.edu Mon Oct 31 11:34:20 2005 From: mdean <@t> uci.edu (mason dean) Date: Mon Oct 31 11:34:33 2005 Subject: [Histonet] calcified cartilage sectioning Message-ID: <728CCF90-189B-438A-B13B-1FC6EF172A53@uci.edu> I'm a graduate student at UC Irvine studying calcification of cartilage in sharks and rays. This type of cartilage has long been equated with mammalian hyaline cartilage, but it's starting to look (from our biochemical and mechanical properties data) like it might not be that simple. Probably the oddest gross morphological feature is that the uncalcified phase (which is similar to hyaline in many ways) is covered by a "bark" of calcified, interlocking tiles. This makes for an abrupt change in material properties! I'm interested in cross-sectioning undecalcified large (~2-4 cm squared) samples, but the more I read the more it seems that may be a pipe dream! Most people are encouraging me to reduce my samples to ~1-2mm, but I would lose an important global understanding of the skeletal element. I'm running out of options, but saw lots of useful histonet postings on hard tissue sectioning and thought I might drop an email. I'd love any ideas/thoughts you have on my options, but am still very much a novice so bear with me! Thanks! Mason ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Mason Dean Ecology & Evolutionary Biology University of California, Irvine 321 Steinhaus Hall; Irvine, CA 92697-2525 ph: 949.824.4830; fax: 949.824.2181 e-mail: mdean@uci.edu website: http://grad.bio.uci.edu/ecoevo/mdean "recuperation is like spring: dormancy + vitality collide." -gretel ehrlich ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ From bliven.laura <@t> marshfieldclinic.org Mon Oct 31 12:16:10 2005 From: bliven.laura <@t> marshfieldclinic.org (bliven.laura@marshfieldclinic.org) Date: Mon Oct 31 12:16:21 2005 Subject: [Histonet] Immunofluorescence - CAP ? Message-ID: <0e1401c5de47$2ba4c4b0$8e0110ac@mfldclinframe.org> A new CAP (College of American Pathologists) question has appeared on our checklist for the next inspection. The question asks, "Are appropriate positive and negative controls performed with each case tested using immunofluorescence?" Sometimes positive controls not abundant and since we are working with only frozen tissue, does anyone have any helpful ideas? Other than past positive cases, is there any other positive tissue that we could use for IgA, IgM, IgG, C3, Fibrinogen, properdin and/or C4? Thanks much, Laura Laura Bliven Histology Lab/IHC Marshfield Laboratories 1000 N. Oak Ave. Marshfield, WI 54449 From TJJ <@t> Stowers-Institute.org Mon Oct 31 12:50:33 2005 From: TJJ <@t> Stowers-Institute.org (Johnson, Teri) Date: Mon Oct 31 12:51:03 2005 Subject: [Histonet] Re: Salivary gland tissue fetal pig problems Message-ID: Dr. Vaughn, My guess is that these pigs were embalmed for study, which means they used embalming fluid of some sort. According to streetdrugs.org, "The percentage of formaldehyde found in embalming fluid ranges anywhere from 5 to 29 percent. The percentage of ethyl alcohol...varies anywhere from 9 to 56 percent." Who knows how they were stored prior to embalming (refrigerated? Slow frozen?) and for how long before the embalming took place. Seems to me this is the most likely cause of the histology you're seeing. You might see if there is a vet school somewhere close that might have the availability of processing freshly fixed samples from necropsy there. Teri Johnson Managing Director Histology Facility Stowers Institute for Medical Research 1000 E. 50th St. Kansas City, MO 64133 From ploykasek <@t> phenopath.com Mon Oct 31 13:45:31 2005 From: ploykasek <@t> phenopath.com (Patti Loykasek) Date: Mon Oct 31 13:45:54 2005 Subject: [Histonet] Immunofluorescence - CAP ? In-Reply-To: <0e1401c5de47$2ba4c4b0$8e0110ac@mfldclinframe.org> Message-ID: Laura, for IgG, IgA, and IgM we use normal tonsil that has been frozen. For the others we use past positive cases. Hope this info helps. Patti Loykasek BS, HTL, QIHC PhenoPath Laboratories Seattle, WA > A new CAP (College of American Pathologists) question has appeared on our > checklist for the next inspection. The question asks, "Are appropriate > positive and negative controls performed with each case tested using > immunofluorescence?" Sometimes positive controls not abundant and since we > are working with only frozen tissue, does anyone have any helpful ideas? Other > than past positive cases, is there any other positive tissue that we could use > for IgA, IgM, IgG, C3, Fibrinogen, properdin and/or C4? > Thanks much, > Laura > > Laura Bliven > Histology Lab/IHC > Marshfield Laboratories > 1000 N. Oak Ave. > Marshfield, WI 54449 > > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------------------------------------------------- This e-mail message, including any attachments, is for the sole use of the intended recipients and may contain privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by e-mail and destroy all copies of the original message, or you may call PhenoPath Laboratories, Seattle, WA U.S.A. at (206) 374-9000. From ajohnson <@t> aipathology.com Mon Oct 31 13:51:36 2005 From: ajohnson <@t> aipathology.com (Amy Johnson) Date: Mon Oct 31 13:57:28 2005 Subject: [Histonet] Ventana Benchmark Message-ID: <016A64931E1ED511B12C0002B3026080523479@SERV001> Here is a question to all those out there that have a Ventana Benchmark. We are going to be inspected by CAP sometime next year. Our lab has gotten the CAP checklist and one of the questions has to do with pH of any of the bulk fluids on the Ventana. Our question to you all is: Do any of you routinely pH the EZ Prep, Reaction buffer and the CC1?? I f you do can you share with us what your ranges are? Thanks Amy Johnson From nancy.troiano <@t> yale.edu Mon Oct 31 14:01:00 2005 From: nancy.troiano <@t> yale.edu (Nancy W. Troiano) Date: Mon Oct 31 14:02:13 2005 Subject: [Histonet] shark cartilage sectioning Message-ID: <5.2.1.1.2.20051031145732.00c24078@email.med.yale.edu> We routinely cut undecalcified rabbit spines 2 -3 cm square embedded in methylmethacrylate (plastic). It is true that the tissue should be smaller for better infiltration and embedding and overall quality of the plastic. You can contact me about our methods via phone or email to discuss further. Nancy W. Troiano, M.S. Associate in Research III Yale University Core Center for Musculoskeletal Research New Haven, CT (203)785-5136 From Virginia.Ross <@t> nrc-cnrc.gc.ca Mon Oct 31 14:05:46 2005 From: Virginia.Ross <@t> nrc-cnrc.gc.ca (Ross, Virginia) Date: Mon Oct 31 14:05:56 2005 Subject: [Histonet] Price for MMA Embedding-Rabbit Femurs Message-ID: > I can't remember how to post on the histonet, can you post this for me > please? > > > > We are wondering if anyone can give us a quote on embedding 48 rabbit > femurs in MMA? How much for 100u sections of those femurs with a > titanium implant? If you have a contact for us we would greatly > appreciate the information. I just want an idea of cost since we have > experienced medical problems with our technicians who handle MMA. We > are in Canada. > Virginia.ross@nrc-cnrc.gc.ca > Virginia Ross > Parathyroid Hormone Group, > Institute for Biological Sciences, > National Research Council > > > From straussj <@t> upstate.edu Mon Oct 31 14:25:13 2005 From: straussj <@t> upstate.edu (Judy Strauss) Date: Mon Oct 31 14:25:48 2005 Subject: Fwd: [Histonet] Price for MMA Embedding-Rabbit Femurs Message-ID: > We are wondering if anyone can give us a quote on embedding 48 rabbit > femurs in MMA? How much for 100u sections of those femurs with a > titanium implant? If you have a contact for us we would greatly > appreciate the information. I just want an idea of cost since we have > experienced medical problems with our technicians who handle MMA. We > are in Canada. > Virginia.ross@nrc-cnrc.gc.ca > Virginia Ross > Parathyroid Hormone Group, > Institute for Biological Sciences, > National Research Council From la.sebree <@t> hosp.wisc.edu Mon Oct 31 14:52:58 2005 From: la.sebree <@t> hosp.wisc.edu (Sebree Linda A.) Date: Mon Oct 31 14:55:26 2005 Subject: [Histonet] Ventana Benchmark Message-ID: We always pH our reaction buffer and it tends to be in the 7.2 to 7.4 range. We don't pH any of the other proprietary buffers, i.e. CC1, CC2, EZ Prep from Ventana. I would consult VMS and see what they say about this subject; we would be interested in knowing as well. I do remember that I questioned them once about pHing the EZ Prep and they said not to because its basically a detergent. Linda A. Sebree University of Wisconsin Hospital & Clinics IHC/ISH Clinical & Research Laboratory DM223-VA 600 Highland Ave. Madison, WI 53792 (608)265-6596 FAX: (608)262-7174 -----Original Message----- From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Amy Johnson Sent: Monday, October 31, 2005 1:52 PM To: Histonet (E-mail) Subject: [Histonet] Ventana Benchmark Here is a question to all those out there that have a Ventana Benchmark. We are going to be inspected by CAP sometime next year. Our lab has gotten the CAP checklist and one of the questions has to do with pH of any of the bulk fluids on the Ventana. Our question to you all is: Do any of you routinely pH the EZ Prep, Reaction buffer and the CC1?? I f you do can you share with us what your ranges are? Thanks Amy Johnson _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet From CBWright <@t> nmcsd.med.navy.mil Mon Oct 31 15:39:55 2005 From: CBWright <@t> nmcsd.med.navy.mil (Wright, Clarissa B CIV) Date: Mon Oct 31 15:38:51 2005 Subject: [Histonet] Ventana Benchmark Message-ID: <4D840F2F36912841BD8B2484E8C6839316C7DFDB@nmc-sdca-exch02> Amy, We only pH the Reaction Buffer. Our readings are 7.6 +/- .2 as stated on the package insert. EZ Prep is detergent, and CC1 lots are pre-diluted and QC'd by Ventana . Kris Wright -----Original Message----- From: Amy Johnson [mailto:ajohnson@aipathology.com] Sent: Monday, October 31, 2005 1:52 PM To: Histonet (E-mail) Subject: [Histonet] Ventana Benchmark Here is a question to all those out there that have a Ventana Benchmark. We are going to be inspected by CAP sometime next year. Our lab has gotten the CAP checklist and one of the questions has to do with pH of any of the bulk fluids on the Ventana. Our question to you all is: Do any of you routinely pH the EZ Prep, Reaction buffer and the CC1?? I f you do can you share with us what your ranges are? Thanks Amy Johnson _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This document may contain information covered under the Privacy Act, 5 USC 552(a), Health Insurance Portability and Accountability Act, Public Law 104-191, and DoD Directive 6025.18. It must be protected in accordance with those provisions. From gcallis <@t> montana.edu Mon Oct 31 15:47:14 2005 From: gcallis <@t> montana.edu (Gayle Callis) Date: Mon Oct 31 15:47:28 2005 Subject: [Histonet] shark cartilage sectioning In-Reply-To: <5.2.1.1.2.20051031145732.00c24078@email.med.yale.edu> References: <5.2.1.1.2.20051031145732.00c24078@email.med.yale.edu> Message-ID: <6.0.0.22.1.20051031144604.01b62f68@gemini.msu.montana.edu> Nance also has a lovely, editor's award publication in J of Histotechnology. Hopefully she has a reprint for you!!! At 01:01 PM 10/31/2005, you wrote: >We routinely cut undecalcified rabbit spines 2 -3 cm square embedded in >methylmethacrylate (plastic). It is true that the tissue should be >smaller for better infiltration and embedding and overall quality of the >plastic. You can contact me about our methods via phone or email to >discuss further. > >Nancy W. Troiano, M.S. >Associate in Research III >Yale University Core Center for Musculoskeletal Research >New Haven, CT > >(203)785-5136 > > >_______________________________________________ >Histonet mailing list >Histonet@lists.utsouthwestern.edu >http://lists.utsouthwestern.edu/mailman/listinfo/histonet Gayle Callis Research Histopathology Supervisor Veterinary Molecular Biology Montana State University - Bozeman PO Box 173610 Bozeman MT 59717-3610 406 994-6367 406 994-4303 (FAX) From vazquezr <@t> ohsu.edu Mon Oct 31 16:28:02 2005 From: vazquezr <@t> ohsu.edu (Robyn Vazquez) Date: Mon Oct 31 16:28:27 2005 Subject: [Histonet] Purcase a microwave tissue processor Message-ID: Try Belair 973-912-8900 or Internation Medical Equipment 800-543-8496 Margaret O'Donnell Robyn OHSU From lrichey <@t> u.washington.edu Mon Oct 31 19:30:46 2005 From: lrichey <@t> u.washington.edu (Lori K. Richey) Date: Mon Oct 31 19:30:55 2005 Subject: [Histonet] Immunofluorescence - CAP ? In-Reply-To: <0e1401c5de47$2ba4c4b0$8e0110ac@mfldclinframe.org> Message-ID: For IgA,M and G we use frozen tonsil. Fibrinogen we use frozen placenta. If its C4d? we use frozen kidney. On Mon, 31 Oct 2005 bliven.laura@marshfieldclinic.org wrote: > > A new CAP (College of American Pathologists) question has appeared on our checklist for the next inspection. The question asks, "Are appropriate positive and negative controls performed with each case tested using immunofluorescence?" Sometimes positive controls not abundant and since we are working with only frozen tissue, does anyone have any helpful ideas? Other than past positive cases, is there any other positive tissue that we could use for IgA, IgM, IgG, C3, Fibrinogen, properdin and/or C4? > Thanks much, > Laura > > Laura Bliven > Histology Lab/IHC > Marshfield Laboratories > 1000 N. Oak Ave. > Marshfield, WI 54449 > > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet >