[Histonet] mouse skin processing for paraffin embedding
mesruh turkekul
turkekul <@t> gmail.com
Fri Aug 10 09:22:51 CDT 2007
Dear All,
I have problems sectioning mouse skin and tail.
My protocol is :
4% PFA 16h at 4 C
wash in PBS
70% Ethanol 30 min RT
85% Ethanol 30 min RT
95% Ethanol 30 min RT
100% Ethanol 2X 30 min each RT
histoclear 3X 30 min each RT
1:1 paraffin-histoclear 45 min at 60C oven
paraffin 3X 30 min each at 60 C oven w/ vacuum.
embedding
I would like to know if skin in particular requires something different in
terms of processing or even sectioning.
Thank you for your valuable opinions.
Cheers,
Mesruh Turkekul
Memorial Sloan-Kettering Cancer Center
New YORK
646 888 2209
On 7/22/07, histonet-request <@t> lists.utsouthwestern.edu <
histonet-request <@t> lists.utsouthwestern.edu> wrote:
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> Today's Topics:
>
> 1. Problems with the Histonet? (Dolores Townsend)
> 2. MWO validation. (Rene J Buesa)
> 3. Shandon Sequenza(r) Immunostaining Center (Igor Deyneko)
> 4. Placenta returns (Bryan Llewellyn)
> 5. Question? (Maribel Santiago)
> 6. paraffin heater (Steven Coakley)
> 7. Looking for Angie Bailey (Victoria Baker)
> 8. No posts (machocraig)
> 9. Listeria By Immunohistochemistry (Marilyn Johnson)
> 10. Re: Placenta returns (Joe Nocito)
> 11. Re: Question? (Joe Nocito)
> 12. RE: Unsuccessful Muscle Lipid Staining (Liz Chlipala)
> 13. RE: paraffin heater (Weems, Joyce)
>
>
> ----------------------------------------------------------------------
>
> Message: 1
> Date: Fri, 20 Jul 2007 11:31:03 -0400
> From: "Dolores Townsend" <TownsendD <@t> childrensdayton.org>
> Subject: [Histonet] Problems with the Histonet?
> To: histonet <@t> lists.utsouthwestern.edu
> Message-ID: <s6a09d10.000 <@t> nw-gwia2>
> Content-Type: text/plain; charset=US-ASCII
>
> I have not received any message from the Histonet since Wednesday
> morning. Anyone else having this problem?
> Dolores
>
>
> ------------------------------
>
> Message: 2
> Date: Fri, 20 Jul 2007 08:45:46 -0700 (PDT)
> From: Rene J Buesa <rjbuesa <@t> yahoo.com>
> Subject: [Histonet] MWO validation.
> To: Patsy Ruegg <pruegg <@t> ihctech.net>,
> histonet <@t> lists.utsouthwestern.edu
> Message-ID: <484779.21679.qm <@t> web61221.mail.yahoo.com>
> Content-Type: text/plain; charset=iso-8859-1
>
> Hi Patsy:
> I completely agree with you. A few days ago I advised an MD do to hold
> using MWO for breast that was going to be used with IHC tests.
> My argument was as follows:
> 1- FDA has approved Her2Neu for FFPE tissues, and they say NOTHING about
> MWO although it could be assumed that it was using conventional processig
> 2-DAKO developed their protocol (later approved by FDA) using conventional
> tissue processors
> 3- IF something happens and a lawsuit is brought, any savy lawyer (and
> they are savy enough) could "dig-out" the type of processing used and IF it
> was MWO it is likely that their argument could be accepted by a jury.
> I don't think that a lab individual validaiton could hold in court
> (against an FDA approved procedure). NEW guidelines by the FDA would be
> needed (as you point out).
> MWO users beware!
> René J.
>
> Patsy Ruegg <pruegg <@t> ihctech.net> wrote:
> Rene,
> Has mw processing been validated for IHC (especially for her2neu) by
> running
> 25-100 cases side by side (one piece conventionally processed and the
> other
> mw processed from each of the 25-100 cases) then running the IHC? Until
> this is done and reported the new guidelines for her2 testing will require
> that any deviation from conventional processing and fixation for 6-48hr
> using the FDA approved her2neu kits, labs using mw processing will not be
> in
> compliance.
> Patsy
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Rene J
> Buesa
> Sent: Tuesday, July 17, 2007 10:17 AM
> To: Weaver, Colin; histonet <@t> lists.utsouthwestern.edu
> Subject: Re: [Histonet] Microwave v conventional processing
>
> Colin:
> Using the adequate protocols MW processing renders equivalent results to
> "conventional" tissue processing, that is the general concensus.
> The thing is that unless you use an automated MW tissue processor, a
> histotech will have to attend to the process and change reagents manually.
> This can lead to 2 problems: higher exposure of the HT to (usually hot)
> chemicals and some degree of inconsistency in the protocol because the
> time
> in each reagent could vary slightly different between runs.
> Consider that a few minutes in a conventional protocol is a much lower
> percentage of the time in the reagent, than the same amount of time in a
> much faster protocol completed with a MW tissue processor.
> MW processing should be an option when TAT is an issue and even then there
> are numerous manual steps independent of the time the tissue is involved
> in
> the processing step; they are independent of the processing technology and
> usually count for the greater part of the total TAT.
> Under separate cover I am sending you an article of mine wher I analyze
> this issue.
> René J.
>
> "Weaver, Colin" wrote:
> Hi - we are trying to go down the microwave route in processing but
> inevitably some of our veterinary pathologists are questioning whether
> microwave sections are as "good" as conventional processing. Can anyone
> point me in the right direction to find any comparison done between
> microwave processing and conventional overnight processing with regard
> to section and staining quality.
>
>
> Colin Weaver
> Veterinary Laboratories Agency (VLA)
>
> This email and any attachments is intended for the named recipient
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> ------------------------------
>
> Message: 3
> Date: Fri, 20 Jul 2007 11:46:06 -0400
> From: "Igor Deyneko" <igor.deyneko <@t> gmail.com>
> Subject: [Histonet] Shandon Sequenza(r) Immunostaining Center
> To: histonet <@t> lists.utsouthwestern.edu
> Message-ID:
> <35e16a770707200846v4c873163v6cbda858caa93950 <@t> mail.gmail.com>
> Content-Type: text/plain; charset=ISO-8859-1; format=flowed
>
> Hello.
> I was just wondering whether or not someone uses Shandon Sequenza(r)
> Immunostaining Center for IHC? The reason i am asking is that do you use
> it
> for all your IHC or in any specific cases. In my experience, I found that
> 2
> slices of tissue on the same slide get different amount of reagents, so
> when
> viewed under the microscope, they light up differently, showing different
> levels of saturation with a chromogen.
> Thank you.
> Igor Deyneko.
> Infinity Pharmaceutical
> Cambridge,MA.
>
>
> ------------------------------
>
> Message: 4
> Date: Fri, 20 Jul 2007 09:42:50 -0700
> From: Bryan Llewellyn <llewllew <@t> shaw.ca>
> Subject: [Histonet] Placenta returns
> To: Histonet <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <000501c7caed$02beb120$ff144246 <@t> yourlk4rlmsu>
> Content-Type: text/plain; format=flowed; charset=iso-8859-1;
> reply-type=original
>
> I thought this might be of interest. We had a discussion on this issue
> several months ago, I believe.
>
>
> http://www.ctv.ca/servlet/ArticleNews/story/CTVNews/20070720/placenta_hospital_070720/20070720?hub=Health
>
> Bryan Llewellyn
>
>
>
>
> ------------------------------
>
> Message: 5
> Date: Fri, 20 Jul 2007 18:45:26 +0000
> From: Maribel Santiago <minniesann <@t> hotmail.com>
> Subject: [Histonet] Question?
> To: <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <BAY117-W27DBE3D420B6F55DD144C8D5F40 <@t> phx.gbl>
> Content-Type: text/plain; charset="iso-8859-1"
>
> What is going on with histonet? I'm not receiving emails since wednesday
> afternoon! Minnie
> _________________________________________________________________
> Express yourself instantly with MSN Messenger! Download today it's FREE!
> http://messenger.msn.click-url.com/go/onm00200471ave/direct/01/
>
> ------------------------------
>
> Message: 6
> Date: Fri, 20 Jul 2007 16:18:10 -0700 (PDT)
> From: Steven Coakley <sjchtascp <@t> yahoo.com>
> Subject: [Histonet] paraffin heater
> To: Histonet <@t> lists.utsouthwestern.edu
> Message-ID: <20070720231810.88168.qmail <@t> web38203.mail.mud.yahoo.com>
> Content-Type: text/plain; charset=iso-8859-1
>
> I'm looking for the maker of a heater used to remove paraffin from the
> sides of paraffin tissue cassettes. I used one a a facility I temped at in
> Milwaukee but didn't get the manufacturer.
>
> Thanks,
>
> Steve
>
>
> ---------------------------------
> Be a better Globetrotter. Get better travel answers from someone who
> knows.
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>
> ------------------------------
>
> Message: 7
> Date: Sat, 21 Jul 2007 10:29:40 -0400
> From: "Victoria Baker" <bakevictoria <@t> gmail.com>
> Subject: [Histonet] Looking for Angie Bailey
> To: "Histo Net list server" <HistoNet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <4f016b690707210729v4e91e191qab17916456c2a662 <@t> mail.gmail.com>
> Content-Type: text/plain; charset=ISO-8859-1; format=flowed
>
> Hi
>
> I am posting this for a friend if anyone know how I can locate Angie
> Bailey, please let me know!
>
> Thanks in advance.
>
> Vikki Baker
>
>
>
> ------------------------------
>
> Message: 8
> Date: Sat, 21 Jul 2007 14:41:39 -0600
> From: "machocraig" <machocraig <@t> hotmail.com>
> Subject: [Histonet] No posts
> To: <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <BAY135-DAV1401862C5C23E5B556D2F9D9F50 <@t> phx.gbl>
> Content-Type: text/plain; format=flowed; charset="Windows-1252";
> reply-type=original
>
> Hi,
> I am not getting any posts.
> Is histonet out of commission?
>
> Craig
>
>
>
> ------------------------------
>
> Message: 9
> Date: Sun, 22 Jul 2007 07:26:03 -0600
> From: "Marilyn Johnson" <marjoh3 <@t> telus.net>
> Subject: [Histonet] Listeria By Immunohistochemistry
> To: <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <000801c7cc63$da331d40$6401a8c0 <@t> VALUED20606295>
> Content-Type: text/plain; charset="iso-8859-1"
>
> Hi Histonetters,
> Thanks to all of you who replied to my request.
> Greatly appreciated.
>
> Marilyn Johnson
> Food Safety Division
> Alberta Agriculture
> Edmonton, AB. Canada
>
> ------------------------------
>
> Message: 10
> Date: Sun, 22 Jul 2007 09:10:42 -0500
> From: "Joe Nocito" <jnocito <@t> satx.rr.com>
> Subject: Re: [Histonet] Placenta returns
> To: "Bryan Llewellyn" <llewllew <@t> shaw.ca>, "Histonet"
> <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <004401c7cc6a$176b0dc0$d49eae18 <@t> yourxhtr8hvc4p>
> Content-Type: text/plain; format=flowed; charset="iso-8859-1";
> reply-type=response
>
> Bryan,
> two things. First, the placenta was frozen and not placed in formalin.
> Every
> hospital I have worked in always placed the placenta in formalin (ok, some
> might have placed 15 mls in the container, maybe 20). The second thing I
> noticed was that Nevada has no law about returning specimens to the owner.
> I've worked at placed were we were being sued by a guy because he
> wanted
> his finger back, 18 months ago. Even though we sent him copies of he
> Texas
> law and MSDS on 10% NBF, he still found a sleazy lawyer to file suit.
> Also, I've had experiences where the patient has requested their
> specimen back because of religious reasons. So here's my point. People who
> want their specimens back because of religious reasons do so up front.
> People who want to place their specimen on the coffee table for a
> conversational piece will request it back as an after thought. Like Billy
> Bob who wanted his finger back. "Hey Bubba, look what I took home from the
> hospital". "No you idiot, it's not chicken liver, it's my hemorrhoids".
> That's my story and I'm sticking to it.
>
> JTT
> ----- Original Message -----
> From: "Bryan Llewellyn" <llewllew <@t> shaw.ca>
> To: "Histonet" <histonet <@t> lists.utsouthwestern.edu>
> Sent: Friday, July 20, 2007 11:42 AM
> Subject: [Histonet] Placenta returns
>
>
> >I thought this might be of interest. We had a discussion on this issue
> >several months ago, I believe.
> >
> >
> http://www.ctv.ca/servlet/ArticleNews/story/CTVNews/20070720/placenta_hospital_070720/20070720?hub=Health
> >
> > Bryan Llewellyn
> >
> > _______________________________________________
> > Histonet mailing list
> > Histonet <@t> lists.utsouthwestern.edu
> > http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
>
>
>
> ------------------------------
>
> Message: 11
> Date: Sun, 22 Jul 2007 09:13:00 -0500
> From: "Joe Nocito" <jnocito <@t> satx.rr.com>
> Subject: Re: [Histonet] Question?
> To: "Maribel Santiago" <minniesann <@t> hotmail.com>,
> <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <005701c7cc6a$69ae9fc0$d49eae18 <@t> yourxhtr8hvc4p>
> Content-Type: text/plain; format=flowed; charset="iso-8859-1";
> reply-type=original
>
> I think the Histonet went on vacation. Hey, it is summer you know. Except
> in
> Texas, it's monsoon season. I picked a real winner of a summer to take
> off.
> Okay, if you must know, I AM walking around with the a big L on my
> forehead.
>
> JTT
> ----- Original Message -----
> From: "Maribel Santiago" <minniesann <@t> hotmail.com>
> To: <histonet <@t> lists.utsouthwestern.edu>
> Sent: Friday, July 20, 2007 1:45 PM
> Subject: [Histonet] Question?
>
>
> What is going on with histonet? I'm not receiving emails since wednesday
> afternoon! Minnie
> _________________________________________________________________
> Express yourself instantly with MSN Messenger! Download today it's FREE!
>
> http://messenger.msn.click-url.com/go/onm00200471ave/direct/01/_______________________________________________
> Histonet mailing list
> Histonet <@t> lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
>
>
>
> ------------------------------
>
> Message: 12
> Date: Sun, 22 Jul 2007 10:13:27 -0600
> From: "Liz Chlipala" <liz <@t> premierlab.com>
> Subject: RE: [Histonet] Unsuccessful Muscle Lipid Staining
> To: "Son Quang Duong" <sqd3f <@t> cms.mail.virginia.edu>,
> <histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <EE33BE5C905A3046A7FF8F58A64C8E4B01A33C <@t> server.PremierLab.local>
> Content-Type: text/plain; charset="windows-1250"
>
> Son
>
> We did this in the past, we post fixed in osmium, it turned out quite
> nice. I'll send images in a different e-mail. We fixed in formalin and
> then post fixed the muscle in osmium overnight, we were working on mouse
> tibialis anterior so we kept the entire muscle intact and processed the
> whole muscle and then cut the muscle in half prior to embedding. We found
> that the unstained sections turned out the best for demonstration of the
> fat. I might have a material and methods some where since I think they
> published on it. I'll send that to you with the images.
>
> Liz
>
>
> Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
> Manager
> Premier Laboratory, LLC
> P.O. Box 18592
> Boulder, CO 80308
> phone (303) 735-5001
> fax (303) 735-3540
> liz <@t> premierlab.com
> www.premierlab.com
>
> Ship to Address:
>
> Premier Laboratory, LLC
> University of Colorado at Boulder
> MCDB, Room A3B40
> Boulder, CO 80309
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:
> histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Sonny Duong
> Sent: Sunday, July 22, 2007 3:31 AM
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] Unsuccessful Muscle Lipid Staining
>
> Hi all,
>
> I have been trying to stain mouse muscle tissue for intramuscular
> triglyceride droplets with Oil Red O (dissolved in triethyl phosphate) for
> some time now, and my results have been disastrous. I've tried it on both
> fresh frozen and formaldehyde fixed tissues (and cryoprotected)
> cryosections, but almost always it seems that the lipid droplets within the
> fibers (maybe?) leak out of the cells and conglomerate on the edges of the
> tissue and within gaps between the cells. Very few of the fibers, if any,
> display any lipid droplet staining and it is always very weak. The lipid
> droplets all conglomerate in roughly the same positions between serial
> sections as well, which leads me to believe something is happening during
> the freezing/fixation process, or the cutting in the cryosection. Does
> anyone have any suggestion as to what I could be doing wrong here (i.ecutting technique, temperature, fixatives)? Also, does anyone have any
> experience with using ORO in triethyl phosphate?
>
> Thank you all for your time,
> Sonny Duong
> University of Virginia
> Green Lab
>
>
>
> _______________________________________________
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>
>
>
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>
> ------------------------------
>
> Message: 13
> Date: Sun, 22 Jul 2007 12:43:11 -0400
> From: "Weems, Joyce" <JWEEMS <@t> sjha.org>
> Subject: RE: [Histonet] paraffin heater
> To: "Steven Coakley" <sjchtascp <@t> yahoo.com>,
> <Histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <1CD6831EB9B26D45B0A3EAA79F7EBD3203FF9EE6 <@t> sjhaexc02.sjha.org>
> Content-Type: text/plain; charset="us-ascii"
>
> ThermoFisher Scientific - formerly
> ThermoElectronFisherScientificThermoShandonShandonLipshawLabVision...
> Etc!
> I would look on the web site. I'm not sure if the product number changed
> after all the merges...
>
> Good luck! j
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Steven
> Coakley
> Sent: Friday, July 20, 2007 7:18 PM
> To: Histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] paraffin heater
>
> I'm looking for the maker of a heater used to remove paraffin from the
> sides of paraffin tissue cassettes. I used one a a facility I temped at
> in Milwaukee but didn't get the manufacturer.
>
> Thanks,
>
> Steve
>
>
> ---------------------------------
> Be a better Globetrotter. Get better travel answers from someone who
> knows.
> Yahoo! Answers - Check it out.
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>
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