[Histonet] Re: Histonet Digest, Vol 42, Issue 33
garret.t.miyamoto <@t> us.army.mil
garret.t.miyamoto <@t> us.army.mil
Wed May 23 15:00:25 CDT 2007
----- Original Message -----
From: histonet-request <@t> lists.utsouthwestern.edu
Date: Wednesday, May 23, 2007 7:13 am
Subject: Histonet Digest, Vol 42, Issue 33
To: histonet <@t> lists.utsouthwestern.edu
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> Today's Topics:
>
> 1. RE: Histonet Digest, Vol 42, Issue 32 (Kobler, James)
> 2. Ethanol ready-to-use for LCM (Castillos, Luminita)
> 3. RE: NK cells in mouse tissue and PLP fixation (Gayle Callis)
> 4. RE: -20C acetone fixative (C.M. van der Loos)
> 5. RE: RE: -20C acetone fixative (Edwards, R.E.)
> 6. Cryomold adapters (Martha Ward)
> 7. Cell Blocks on small buttons... (Lott, Robert)
> 8. celloidin-ether/alc (Mary Lou Norman)
> 9. Work hours (Patti Loykasek)
> 10. RE: celloidin-ether/alc (Rittman, Barry R)
> 11. Re: Work hours (Rene J Buesa)
> 12. RE: Work hours (Mike Pence)
>
>
> -------------------------------------------------------------------
> ---
>
> Message: 1
> Date: Wed, 23 May 2007 10:20:34 -0400
> From: "Kobler, James" <JKOBLER <@t> PARTNERS.ORG>
> Subject: [Histonet] RE: Histonet Digest, Vol 42, Issue 32
> To: <histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <6D1DFC2837CEAE4BBBDED596071430854D07D4 <@t> PHSXMB4.partners.org>
> Content-Type: text/plain; charset="us-ascii"
>
> Dear histonet contributors,
>
> I would greatly appreciate any advice about antibodies that work
> well with
> ferret tissue (and any related processing tips). We are
> particularly interested
> in antibodies to markers for fibroblasts, myofibroblasts,
> endothelial, smooth
> muscle and inflammatory cells, as well as extracellular matrix.
> Thanks very
> much,
>
> Jim Kobler, Massachusetts General Hospital
>
>
>
>
>
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> ------------------------------
>
> Message: 2
> Date: Wed, 23 May 2007 10:28:19 -0400
> From: "Castillos, Luminita" <lcastillos <@t> cogenics.com>
> Subject: [Histonet] Ethanol ready-to-use for LCM
> To: <histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <DE517009A0E1B345BAE029F86CADECCB059934 <@t> ncmsx2.gpi.genaissance.com>
> Content-Type: text/plain; charset="US-ASCII"
>
>
> Hi,
>
> Does any know from where to buy ready-to-use ethanol (100%, 95%, 70%)
> molecular biology grade and RNase-free, compatible for Laser Capture
> microdissection staining protocol?. Thank you very much.
>
> L,
>
>
> ------------------------------
>
> Message: 3
> Date: Wed, 23 May 2007 08:43:03 -0600
> From: Gayle Callis <gcallis <@t> montana.edu>
> Subject: RE: [Histonet] NK cells in mouse tissue and PLP fixation
> To: Bruce Abaloz <brucea <@t> unimelb.edu.au>,
> Histonet <@t> lists.utsouthwestern.edu
> Message-ID:
> <6.0.0.22.1.20070523083057.01b3a148 <@t> gemini.msu.montana.edu>
> Content-Type: text/plain; charset="us-ascii"; format=flowed
>
> Sonya,
>
> You can try PLP immersion fixation followed by sucrose
> cryoprotection, but
> the publication I read (reference was in a Histonet message
> yesterday) did
> a fixation study with PLP and other fixatives. They had little
> success
> with the NK1.1 (P136) antibody following immersion fixation with
> all
> fixatives, before snap freezing or after fresh tissue frozen
> sections were
> cut. They had nice staining following PERFUSION (with PLP) of the
> mouse. After perfusion, and once the tissues are dissected out,
> then
> immersion into this fixative should ensure complete fixation. You
> may want
> to do final fixation in PLP overnight, some do it for 5 - 6 hours
> longer
> here, then do the 30% sucrose cryoprotection at 4C. Just remove
> the tissue
> from fixative and go to the sucrose solution as Bruce suggested.
> Small
> fixed tissues often float on top of the sucrose solution. A
> general rule
> of thumb the that cryoprotection is done, tissues sink to the
> bottom of the
> container.
>
> Some people use a sucrose gradient, often seen in the literature,
> but we
> have never found that necessary. We blot off the excess sucrose
> solution
> before embedding in OCT.
>
> Hopefully you have access to a perfusion methods IF your immersion
> fails.
> Gayle Callis HTL, HT, MT(ASCP)
> Research Histopathology Supervisor
> Veterinary Molecular Biology
> Montana State University
> Bozeman MT 59717
>
>
> At 06:12 PM 5/22/2007, you wrote:
> >I just snap frooze the fresh tissue in isopentane on dry ice -
> this has
> >worked with all the other markers I've been looking at but I
> gather NK
> >cells are more tricky.
> >I want to try PLP fixation before I freeze so do I put it
> straight from
> >PLP to 30% sucrose or do I wash/use increasing amounts of sucrose.
> >Once I've left the tissue overnight in sucrose can I then freeze
> it in
> >TissueTek on dry ice as normal?
> >
> >Thanks
> >Sonya
> >
> >-----Original Message-----
> >From: Gayle Callis [mailto:gcallis <@t> montana.edu]
> >Sent: 22 May 2007 15:49
> >To: Martin S.
> >Subject: Re: [Histonet] NK cells in mouse tissue and PLP fixation
> >
> >How did you fix in the first place?
> >
> >You need to sucrose cryoprotect after PLP fixation, in 30% sucrose
> >overnight BEFORE snap freezing the tissues.
> >
> >At 04:01 AM 5/22/2007, you wrote:
> >>Has anyone done any staining for NK cells in mouse tissue. We
> have an
> >>antibody against NK1.1 (PK136) but it doesnt seem to work on
> >>fresh-frozen tissues. I have looked through the literature and some
> >>people seem to have got iot to work on fresh-frozens while
> others have
> >>used PLP fixation before embedding in Tissue Tek.
> >>
> >>Has anyone had any experience with this Ab?
> >>
> >>Also, if I fix the tissues with PLP before freezing then how
> should I
> >>treat them before outting them in TissueTek to freeze?
> >>
> >>Thanks
> >>Sonya
> >>
> >>
> >>
> >>_______________________________________________
> >>Histonet mailing list
> >>Histonet <@t> lists.utsouthwestern.edu
> >>http://lists.utsouthwestern.edu/mailman/listinfo/histonet
> >
> >Gayle Callis
> >MT,HT,HTL(ASCP)
> >Research Histopathology Supervisor
> >Veterinary Molecular Biology
> >Montana State University - Bozeman
> >PO Box 173610
> >Bozeman MT 59717-3610
> >
> >
> >
> >
> >
> >
> >
> >
> >_______________________________________________
> >Histonet mailing list
> >Histonet <@t> lists.utsouthwestern.edu
> >http://lists.utsouthwestern.edu/mailman/listinfo/histonet
> >--
> >BRUCE ABALOZ
> >HISTOLOGIST PH:61383446282
> >DEPT. of ZOOLOGY EMAIL: brucea <@t> unimelb.edu.au
> >THE UNIVERSITY of MELBOURNE. FAX:61383447909
> >VICTORIA. AUSTRALIA 3010
> > Nobody Can Make You Feel Inferior Without YOUR
> Permission -
> > Eleanor Roosevelt
> > WHETHER YOU THINK YOU CAN-OR WHETHER YOU
> THINK YOU
> > CAN'T-YOU'RE RIGHT!!
> > Be reasonable...
> Demand the
> > impossible.....
> > <')))>>< <')))>><
> > <')))>>< <')))>><
> > P Please consider the environment before printing
> this e-mail.
> >
> >_______________________________________________
> >Histonet mailing list
> >Histonet <@t> lists.utsouthwestern.edu
> >http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
>
>
>
> ------------------------------
>
> Message: 4
> Date: Wed, 23 May 2007 16:48:47 +0200
> From: "C.M. van der Loos" <c.m.vanderloos <@t> amc.uva.nl>
> Subject: [Histonet] RE: -20C acetone fixative
> To: histonet <@t> lists.utsouthwestern.edu
> Message-ID: <37e62d383380.38338037e62d <@t> amc.uva.nl>
> Content-Type: text/plain; charset="us-ascii"
>
>
> Long time ago they told me it is a milder fixative at -20C
> than at RT
> ???
>
> Chris van der Loos, PhD
> Dept. of Pathology
> Academic Medical Center M2-230
> Meibergdreef 9
> NL-1105 AZ Amsterdam
> The Netherlands
>
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [[1]mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On
> Behalf Of
> Charles,
> Roger
> Sent: Tuesday, May 22, 2007 12:29 PM
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] -20C acetone fixative
> Does any know or remember why acetone, when used as a fixative, is
> always used "ice cold" or in our case at -20C?
> Roger Charles
> Microbiologist
> Pennsylvania Veterinary Laboratory
> 2305 N Cameron St
> Harrisburg, PA 17110
> 717-787-8808
>
> References
>
> 1. javascript:main.compose('new', 't=histonet-
> bounces <@t> lists.utsouthwestern.edu')
>
> ------------------------------
>
> Message: 5
> Date: Wed, 23 May 2007 16:15:14 +0100
> From: "Edwards, R.E." <ree3 <@t> leicester.ac.uk>
> Subject: RE: [Histonet] RE: -20C acetone fixative
> To: "C.M. van der Loos" <c.m.vanderloos <@t> amc.uva.nl>,
> <histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <DC88BEDFD1FC3F468D0376A7C75465F7101C6E24 <@t> Saffron.cfs.le.ac.uk>
> Content-Type: text/plain; charset="US-ASCII"
>
> And me, in particular for enzyme histochemistry...
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of
> C.M. van
> der Loos
> Sent: 23 May 2007 15:49
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] RE: -20C acetone fixative
>
>
> Long time ago they told me it is a milder fixative at -20C
> than at
> RT
> ???
>
> Chris van der Loos, PhD
> Dept. of Pathology
> Academic Medical Center M2-230
> Meibergdreef 9
> NL-1105 AZ Amsterdam
> The Netherlands
>
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [[1]mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf
> Of
> Charles,
> Roger
> Sent: Tuesday, May 22, 2007 12:29 PM
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] -20C acetone fixative
> Does any know or remember why acetone, when used as a fixative, is
> always used "ice cold" or in our case at -20C?
> Roger Charles
> Microbiologist
> Pennsylvania Veterinary Laboratory
> 2305 N Cameron St
> Harrisburg, PA 17110
> 717-787-8808
>
> References
>
> 1. javascript:main.compose('new',
> 't=histonet-bounces <@t> lists.utsouthwestern.edu')
> _______________________________________________
> Histonet mailing list
> Histonet <@t> lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
>
>
> ------------------------------
>
> Message: 6
> Date: Wed, 23 May 2007 11:02:18 -0400
> From: "Martha Ward" <mward <@t> wfubmc.edu>
> Subject: [Histonet] Cryomold adapters
> To: <histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <61135F0455D33347B5AAE209B903A3041B1C9EBD <@t> EXCHVS2.medctr.ad.wfubmc.edu>
>
> Content-Type: text/plain; charset="us-ascii"
>
> I am looking for cryomold adapters and I hope someone can help. The
> ones we are using have a round center and are a white, hard
> plastic. I
> can find adapters that have a square center but that isn't what we
> want.I have tried the usual vendors but was hoping someone could
> help me.
> Thanks in advance for your help.
>
> Martha Ward, MT (ASCP) QIHC
> Assistant Manager, Molecular Diagnostics Lab
> Wake Forest University Baptist Medical Center
> Medical Center Blvd.
> Winston-Salem, NC 27157
> 336-716-2756
> mward <@t> wfubmc.edu
>
>
>
> ------------------------------
>
> Message: 7
> Date: Wed, 23 May 2007 10:23:14 -0500
> From: "Lott, Robert" <Robert.Lott <@t> TriadHospitals.com>
> Subject: [Histonet] Cell Blocks on small buttons...
> To: <histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <673832E27C45FC4D97EF758FFC777C27020E03DC <@t> CPRTEVS01.triadhospitals.net>
>
> Content-Type: text/plain; charset="us-ascii"
>
> Hi Everyone,
>
> We are interested in hearing about different procedures for recovering
> small cytology aspirates (any fluid really!) ... and making a cell
> blockfrom them using some type of coagulative process/fluid/etc.
> ... so that
> virtually all of the cells are retained!
>
>
>
> There may be some commercial products available ... would like to hear
> about those as well.!!
>
>
>
> Thanks!!!!
>
>
>
> Robert
>
>
>
> Robert L. Lott, HTL(ASCP)
>
> Manager, Anatomic Pathology
>
> LabFirst / Trinity Medical Center - formerly
>
> Montclair Baptist Medical Center
>
> 800 Montclair Road
>
> Birmingham, AL 35213
>
> 205-592-5388 phone
>
> 205-592-5646 fax
>
> robert.lott <@t> triadhospitals.com
>
>
>
>
>
>
> ------------------------------
>
> Message: 8
> Date: Wed, 23 May 2007 11:26:13 -0400
> From: Mary Lou Norman <mlm11 <@t> cornell.edu>
> Subject: [Histonet] celloidin-ether/alc
> To: histonet <@t> lists.utsouthwestern.edu
> Message-ID:
> <6.2.1.2.2.20070523111029.04899930 <@t> postoffice9.mail.cornell.edu>
> Content-Type: text/plain; charset="us-ascii"; format=flowed
>
> Hello Histonet,
>
> Is there any other choice besides the cellodin-ether/alcohol
> coating? My
> parloidin is taking 'forever' to dissolve -- I have read a week
> but I could
> have used it a week ago.
>
> Thanks for your help.
> Mary Lou Norman
>
>
>
>
> ------------------------------
>
> Message: 9
> Date: Wed, 23 May 2007 08:42:22 -0700
> From: Patti Loykasek <ploykasek <@t> phenopath.com>
> Subject: [Histonet] Work hours
> To: histonet <histonet <@t> pathology.swmed.edu>
> Message-ID: <C279AEEE.FD8D%ploykasek <@t> phenopath.com>
> Content-Type: text/plain; charset="US-ASCII"
>
> I was wondering if anyone in histoland has experienced moving from a
> schedule of standard 8 hour day/5 days a week to a 10 hour day/4
> days a
> week with techs having a day off during the week. If so, what are
> the pros
> and cons of this type of schedule. I'm interested in feedback from
> bothbench techs and supervisor/managers. We are in the initial
> stages of
> contemplating this and would appreciate info from anyone that has
> tried it.
> Thank you.
>
>
> 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.
>
>
>
>
> ------------------------------
>
> Message: 10
> Date: Wed, 23 May 2007 11:37:45 -0500
> From: "Rittman, Barry R" <Barry.R.Rittman <@t> uth.tmc.edu>
> Subject: RE: [Histonet] celloidin-ether/alc
> To: "Mary Lou Norman" <mlm11 <@t> cornell.edu>,
> <histonet <@t> lists.utsouthwestern.edu>
> Message-ID:
> <EA1FDD2A141B7448B4B1AFFFCAC08DE406169F41 <@t> UTHEVS1.mail.uthouston.edu>
> Content-Type: text/plain; charset="iso-8859-1"
>
> Mary Lou
> Will dissolve much more rapidly if soak the strips first in
> absolute ethanol for a day o so. The add the ether.
> Barry
>
> ________________________________
>
> From: histonet-bounces <@t> lists.utsouthwestern.edu on behalf of Mary
> Lou Norman
> Sent: Wed 5/23/2007 10:26 AM
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] celloidin-ether/alc
>
>
>
> Hello Histonet,
>
> Is there any other choice besides the cellodin-ether/alcohol
> coating? My
> parloidin is taking 'forever' to dissolve -- I have read a week
> but I could
> have used it a week ago.
>
> Thanks for your help.
> Mary Lou Norman
>
>
> _______________________________________________
> Histonet mailing list
> Histonet <@t> lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
>
>
>
>
> ------------------------------
>
> Message: 11
> Date: Wed, 23 May 2007 09:52:11 -0700 (PDT)
> From: Rene J Buesa <rjbuesa <@t> yahoo.com>
> Subject: Re: [Histonet] Work hours
> To: Patti Loykasek <ploykasek <@t> phenopath.com>, histonet
> <histonet <@t> pathology.swmed.edu>
> Message-ID: <990345.12651.qm <@t> web61216.mail.yahoo.com>
> Content-Type: text/plain; charset=iso-8859-1
>
> I have tried it.
> Cons: If yu don't have a flow of specimens in 4 days equivalent
> to the flow on 5 days, the personnel will be idling some time each
> day. If specimens or workload is available everybody will have
> work to do.
> Pros: it is fantastic having 3 days off weekly. Everybody likes
> that! Ren� J.
>
> Patti Loykasek <ploykasek <@t> phenopath.com> wrote:
> I was wondering if anyone in histoland has experienced moving
> from a
> schedule of standard 8 hour day/5 days a week to a 10 hour day/4
> days a
> week with techs having a day off during the week. If so, what are
> the pros
> and cons of this type of schedule. I'm interested in feedback from
> bothbench techs and supervisor/managers. We are in the initial
> stages of
> contemplating this and would appreciate info from anyone that has
> tried it.
> Thank you.
>
>
> 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.
>
>
> _______________________________________________
> Histonet mailing list
> Histonet <@t> lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
>
>
>
>
> ---------------------------------
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> ------------------------------
>
> Message: 12
> Date: Wed, 23 May 2007 11:58:01 -0500
> From: "Mike Pence" <mpence <@t> grhs.net>
> Subject: RE: [Histonet] Work hours
> To: "Rene J Buesa" <rjbuesa <@t> yahoo.com>, "Patti Loykasek"
> <ploykasek <@t> phenopath.com>, "histonet" <histonet <@t> pathology.swmed.edu>
> Message-ID: <661949901A768E4F9CC16D8AF8F2838CA1C5E3 <@t> IS-E2K3.grhs.net>
> Content-Type: text/plain; charset="iso-8859-1"
>
> How do you decide who gets which days off! Are they floating days?
>
> Mike
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-
> bounces <@t> lists.utsouthwestern.edu] On Behalf Of Rene J Buesa
> Sent: Wednesday, May 23, 2007 11:52 AM
> To: Patti Loykasek; histonet
> Subject: Re: [Histonet] Work hours
>
>
> I have tried it.
> Cons: If yu don't have a flow of specimens in 4 days equivalent
> to the flow on 5 days, the personnel will be idling some time each
> day. If specimens or workload is available everybody will have
> work to do.
> Pros: it is fantastic having 3 days off weekly. Everybody likes
> that! Ren� J.
>
> Patti Loykasek <ploykasek <@t> phenopath.com> wrote:
> I was wondering if anyone in histoland has experienced moving
> from a schedule of standard 8 hour day/5 days a week to a 10 hour
> day/4 days a week with techs having a day off during the week. If
> so, what are the pros and cons of this type of schedule. I'm
> interested in feedback from both bench techs and
> supervisor/managers. We are in the initial stages of contemplating
> this and would appreciate info from anyone that has tried it.
> Thank you.
>
>
> 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.
>
>
> _______________________________________________
> Histonet mailing list
> Histonet <@t> lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
>
>
>
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>
>
>
> ------------------------------
>
> _______________________________________________
> Histonet mailing list
> Histonet <@t> lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
> End of Histonet Digest, Vol 42, Issue 33
> ****************************************
> Re: cell blocks on small buttons
Robert,
I asked someone in our cytology section about your inquiry. What they do is spin down the specimen in a centrifuge, decant the liquid, add about four drops of "HistoGel" (specimen processing gel from Richard-Allen Scientific, 12 vials - 10 ml per vial, reorder number HG 4000-012), respin the specimen and collect the gel button, put it into a tissue specimen bag before placing into the cassette. I hope this will work for you.
Garret
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