[Histonet] RE: Anyone use glass knifes anymore?
I make and use glass knifes for plastic sectioning at 3 microns. I usually have to go through a whole glass strip to get one good knife. Does anyone make them anymore and have the same problem?
~Stephanie Brusig
Weyerhaeuser Company
Propagation of High Value Trees
32901 Weyerhaeuser Way S
WTC-1B10
Federal Way, WA 98001
253.924.6518
-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of histonet-request@lists.utsouthwestern.edu
Sent: Wednesday, May 03, 2006 10:10 AM
To: histonet@lists.utsouthwestern.edu
Subject: Histonet Digest, Vol 30, Issue 3
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Today's Topics:
1. CONTROL BLOCKS NEEDED (Nymeyer, Heather)
2. Large sections (Patsy Ruegg)
3. Autostainer tubes (Patti Loykasek)
4. NSH Active Members (Carrie Diamond)
5. Question (Nita Searcy)
6. Re: Question (Victoria Baker)
7. Re: Large sections (Gayle Callis)
8. RE: Question (Allen, Rhonda)
9. RE: Question (Bartlett, Jeanine (CDC/NCID/VR))
10. Re: Autostainer tubes (Joanne Mauger)
11. Re: POC for Genetic Studies (histology@gradymem.org)
12. antibodies to eIF2 alpha and PPAR gamma (Sarka Lhotak)
13. IHC on frozens (Sarka Lhotak)
14. Fwd: [Histonet] CD Abs (Linresearch@aol.com)
15. gallocyanin stain for nissel (Elizabeth Chlipala)
16. RANK, ER and PR in mouse (Randolph-Habecker, Julie)
17. NADH staining (Yak-Nam Wang)
18. Re: gallocyanine stain for Nissl (John Kiernan)
19. Inflamatory markers (CRAIG BARLOW)
20. Re- processing fatty tissues (Malam Jacqueline)
21. The end of an era (McGovern, Kevin)
22. Mycobacterium bovis (BCG) antibody (Martha Ward)
23. Glycerol cryoprotected tissue and cryostat sectioning
(Kathie A Berghorn)
24. RE: IHC on frozens (Guillermo Palao)
25. Save These Dates For The Pennsylvania State Histology Meeting
(Pamela Marcum)
26. RE: Mycobacterium bovis (BCG) antibody (Elizabeth Chlipala)
----------------------------------------------------------------------
Message: 1
Date: Tue, 2 May 2006 10:03:23 -0700
From: "Nymeyer, Heather"
Subject: [Histonet] CONTROL BLOCKS NEEDED
To:
Message-ID:
<463911DE3F9F8D4AA3EC67632386BC62D1509E@dc1serv78.interiorhealth.ca>
Content-Type: text/plain; charset="us-ascii"
We are in need of the following control blocks
- Leprosy bacilli
- Spirochete
Your assistance is appreciated and thank you in advance.
Heather D. Nymeyer, RT, CEBT
Charge Technologist, Anatomic Pathology
Royal Inland Hospital,
Kamloops, BC.
250-314-2664
------------------------------
Message: 2
Date: Tue, 2 May 2006 10:52:50 -0600
From: "Patsy Ruegg"
Subject: [Histonet] Large sections
To:
Message-ID: <200605021652.k42Gqoog014137@chip.viawest.net>
Content-Type: text/plain; charset="US-ASCII"
Can anyone suggest a resource for getting frozen sections of Elk Larynx?
One of my clients asks:
"I study elk sound production. As one question we investigate the anatomical structure of the larynx. For this purpose I need cross sections of several larynges (approx. 15 larynges). Cross sections at three positions are necessary for each larynx,. The larynges are most likely calcified at varying degrees. I have stored the larynges frozen in saline solution."
Patsy Ruegg, HT(ASCP)QIHC
IHCtech, LLC
Fitzsimmons BioScience Park
12635 Montview Blvd. Suite 215
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: 3
Date: Tue, 02 May 2006 10:31:31 -0700
From: Patti Loykasek
Subject: [Histonet] Autostainer tubes
To: histonet
Message-ID:
Content-Type: text/plain; charset="US-ASCII"
Opps - didn't put in there a supplier other than Dako. We had a backorder problem with Dako on these. Actually turned out to be a glitch in their system, so all is well. Thanks for all of the good info, though. Histotechs are a nice bunch of people!
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: 4
Date: Tue, 2 May 2006 13:37:13 -0400
From: "Carrie Diamond"
Subject: [Histonet] NSH Active Members
To:
Message-ID:
Content-Type: text/plain; charset="us-ascii"
NSH Active Members:
Earlier this week you received a ballot and candidate information for the NSH 2006 Election. Unfortunately, an error occurred at the mail house. The mailing lists were incorrectly matched to the wrong region director information. It lists the incorrect region for which you are eligible to vote. We apologize for this mistake and have mailed a corrected ballot indicating the region director who is running for the position within your region. The ballot has been printed on cream colored paper and will be the only ballot counted from your region. We do hope you will return your ballot at your earliest convenience to the Nominations-Election Committee.
Carrie Diamond
Executive Director
National Society for Histotechnology
4201 Northview Drive, Suite 502
Bowie, MD 20716
P: 301-262-6221
E: carrie@nsh.org
Join us for the
32nd Annual Symposium Convention
Phoenix, AZ
September 8-13, 2006
Registration Details will be available in March
------------------------------
Message: 5
Date: Tue, 02 May 2006 12:49:01 -0500
From: "Nita Searcy"
Subject: [Histonet] Question
To:
Message-ID:
Content-Type: text/plain; charset=US-ASCII
Anyone know of an inexpensive tool for cutting "fresh" 1 mm sections?
Researcher wants to know.
Thanks
------------------------------
Message: 6
Date: Tue, 2 May 2006 11:03:59 -0700 (PDT)
From: Victoria Baker
Subject: Re: [Histonet] Question
To: Nita Searcy ,
histonet@lists.utsouthwestern.edu
Message-ID: <20060502180359.42479.qmail@web52513.mail.yahoo.com>
Content-Type: text/plain; charset=iso-8859-1
The only thing I can think of is what literally looks like an "egg slicer" I think we had ordered it from MOPEC or one of the other surgical/autopsy companies some 6 years ago. It's a metal egg shaped base, and using hi-profile microtome blades you could slice fresh soft tissue (like brain)in thin even pieces. I haven't seen one in sometime, but maybe someone who reads my e-mail will know where they can be purchased these days. If I remember correctly back then it cost about $100.
Good luck
Vikki
--- Nita Searcy wrote:
> Anyone know of an inexpensive tool for cutting "fresh" 1 mm sections?
> Researcher wants to know.
>
> Thanks
>
> _______________________________________________
> Histonet mailing list
> Histonet@lists.utsouthwestern.edu
>
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
__________________________________________________
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Tired of spam? Yahoo! Mail has the best spam protection around http://mail.yahoo.com
------------------------------
Message: 7
Date: Tue, 02 May 2006 12:31:46 -0600
From: Gayle Callis
Subject: Re: [Histonet] Large sections
To: "Patsy Ruegg" ,
Histonet@lists.utsouthwestern.edu
Message-ID:
<6.0.0.22.1.20060502122207.01b50218@gemini.msu.montana.edu>
Content-Type: text/plain; charset="us-ascii"; format=flowed
I don't think the larynx will be calcified, but it will contain both
hyaline and elastic cartilage which could be pretty tough to
cryosection. The Instrumedics should be able to do the job with a heavy
duty knife, maybe even the TC used for undecalcified bone sections but if
the samples are huge, it would require the macro system.
I may be much easier to do paraffin sections and NOT cryosections. The
samples would have to be thawed, simply drop into formalin, fix and process
with a longer schedule to ensure good dehydration, clearing, but most of
all infiltration of paraffin into the cartilage.
At 10:52 AM 5/2/2006, you wrote:
>Can anyone suggest a resource for getting frozen sections of Elk Larynx?
>
>One of my clients asks:
>"I study elk sound production. As one question we investigate the anatomical
>structure of the larynx. For this purpose I need cross sections of several
>larynges (approx. 15 larynges). Cross sections at three positions are
>necessary for each larynx,. The larynges are most likely calcified at
>varying degrees. I have stored the larynges frozen in saline solution."
>
>Patsy Ruegg, HT(ASCP)QIHC
>IHCtech, LLC
>Fitzsimmons BioScience Park
>12635 Montview Blvd. Suite 215
>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
MT,HT,HTL(ASCP)
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: 8
Date: Tue, 2 May 2006 13:32:41 -0500
From: "Allen, Rhonda"
Subject: RE: [Histonet] Question
To: "Victoria Baker" , "Nita Searcy"
,
Message-ID:
Content-Type: text/plain; charset="us-ascii"
There is a brain mold available from EMS. Look under Matrices and they
have different kinds of matrices.
Rhonda Allen BA HT(ASCP)HTL, QIHC
Histotechnology Specialist II
Stowers Institute
1000 E. 50th Street
Kansas City, MO 64110
816-926-4305
-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu
[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Victoria
Baker
Sent: Tuesday, May 02, 2006 1:04 PM
To: Nita Searcy; histonet@lists.utsouthwestern.edu
Subject: Re: [Histonet] Question
The only thing I can think of is what literally looks
like an "egg slicer" I think we had ordered it from
MOPEC or one of the other surgical/autopsy companies
some 6 years ago. It's a metal egg shaped base, and
using hi-profile microtome blades you could slice
fresh soft tissue (like brain)in thin even pieces. I
haven't seen one in sometime, but maybe someone who
reads my e-mail will know where they can be purchased
these days. If I remember correctly back then it cost
about $100.
Good luck
Vikki
--- Nita Searcy wrote:
> Anyone know of an inexpensive tool for cutting
> "fresh" 1 mm sections?
> Researcher wants to know.
>
> Thanks
>
> _______________________________________________
> Histonet mailing list
> Histonet@lists.utsouthwestern.edu
>
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
__________________________________________________
Do You Yahoo!?
Tired of spam? Yahoo! Mail has the best spam protection around
http://mail.yahoo.com
_______________________________________________
Histonet mailing list
Histonet@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
------------------------------
Message: 9
Date: Tue, 2 May 2006 14:24:20 -0400
From: "Bartlett, Jeanine \(CDC/NCID/VR\)"
Subject: RE: [Histonet] Question
To: "Nita Searcy" ,
Message-ID:
Content-Type: text/plain; charset="us-ascii"
Check with Sakura regarding their grossing tools.
Jeanine Bartlett, BS, HT(ASCP)
Centers for Disease Control and Prevention
1600 Clifton Road, MS/G-32
18/SB-114
Atlanta, GA 30333
(404) 639-3590
jeanine.bartlett@cdc.hhs.gov
-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu
[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Nita
Searcy
Sent: Tuesday, May 02, 2006 1:49 PM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Question
Anyone know of an inexpensive tool for cutting "fresh" 1 mm sections?
Researcher wants to know.
Thanks
_______________________________________________
Histonet mailing list
Histonet@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
------------------------------
Message: 10
Date: Tue, 02 May 2006 14:42:30 -0400
From: "Joanne Mauger"
Subject: Re: [Histonet] Autostainer tubes
To: ,
Message-ID:
Content-Type: text/plain; charset=US-ASCII
Patti, Labvision sells the identical vials, in fact I was told they make
the autostainer for Dako-
www.labvision.com
Jo
>>> Patti Loykasek 05/02/06 1:31 PM >>>
Opps - didn't put in there a supplier other than Dako. We had a
backorder
problem with Dako on these. Actually turned out to be a glitch in
their
system, so all is well. Thanks for all of the good info, though.
Histotechs
are a nice bunch of people!
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@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
------------------------------
Message: 11
Date: Tue, 02 May 2006 14:03:38 -0500
From: histology@gradymem.org
Subject: Re: [Histonet] POC for Genetic Studies
To: Laurie Colbert
Cc: histonet@lists.utsouthwestern.edu
Message-ID: <52c3d553056d.53056d52c3d5@onenet.net>
Content-Type: text/plain; charset=us-ascii
The POC goes through Pathology here and we send it out.
Angie Barnett, HTL(ASCP)
Grady Memorial Hospital
Pathology Department
405/224-2258
histology@gradymem.org
----- Original Message -----
From: Laurie Colbert
Date: Thursday, April 27, 2006 1:34 pm
Subject: [Histonet] POC for Genetic Studies
> This question is for those of you who work in hospitals and send
> out POC tissue for genetic studies/chromosome analysis.
>
> Does the tissue get sent out directly from Labor and Delivery, or
> does the tissue go through Pathology and Pathology sends it out?
>
>
>
> _______________________________________________
> Histonet mailing list
> Histonet@lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>
------------------------------
Message: 12
Date: Tue, 02 May 2006 16:17:32 -0400
From: "Sarka Lhotak"
Subject: [Histonet] antibodies to eIF2 alpha and PPAR gamma
To: histonet@lists.utsouthwestern.edu
Message-ID:
Content-Type: text/plain; charset="ISO-8859-1"
Hello Netters,
I am looking for antibodies to phosphorylated eIF2-alpha and PPAR-gamma
that work on mouse tissue, FFPE preferentially. I have tried some with
all kinds of conditions and retrievals with no success. Is anybody
doing this?
Thanks a lot,
desperately,
Sarka Lhotak
McMaster University
Hamilton, Ontario
------------------------------
Message: 13
Date: Tue, 02 May 2006 16:28:32 -0400
From: "Sarka Lhotak"
Subject: [Histonet] IHC on frozens
To: histonet@lists.utsouthwestern.edu
Message-ID:
Content-Type: text/plain; charset="ISO-8859-1"
Hello Netters,
I am trying to get some antibodies work on mouse tissue (see my
previous posting). Since I have not been successful on paraffin
sections I now turned to frozens where I have no experience. I am
getting the same staining (or background of course) on positive and
negative slides (antibody omitted). Even antibodies that I know work
very well on paraffin (my positive control), give me no specific
staining and a background mainly on connective tissue. I've searched
for a proper fixing, drying, dipping in water etc. procedures prior to
staining. I found it very confusing, people swear by different, even
opposing protocols.
So I have a few questions:
What is the best way to prepare/store your sections prior to staining?
Is quenching peroxidase and blocking the same as on paraffin?
Generally, are the primary and secondary antibody concentrations the
same as on paraffin?
Is the MOM problem worse on frozens than on paraffin?
All suggestions are welcome.
Desperately,
Sarka Lhotak
McMaster University
Hamilton, Ontario
------------------------------
Message: 14
Date: Tue, 2 May 2006 18:14:17 EDT
From: Linresearch@aol.com
Subject: Fwd: [Histonet] CD Abs
To: histonet@pathology.swmed.edu
Message-ID: <241.a7d80e3.318933b9@aol.com>
Content-Type: text/plain; charset="us-ascii"
------------------------------
Message: 15
Date: Tue, 2 May 2006 16:31:31 -0600
From: "Elizabeth Chlipala"
Subject: [Histonet] gallocyanin stain for nissel
To:
Message-ID: <000a01c66e38$29664530$0300a8c0@Chlipala>
Content-Type: text/plain; charset="US-ASCII"
I need to perform a gallocyanin stain for nissel on mouse lumbar spinal
cord. I have a protocol from bancroft and it says to stain for 18-48
hours. Can anyone out there give me any pointers on this stain. I also
need to order the gallocyanin and and the only reagent I see is
gallocyanine is that the same thing?
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
------------------------------
Message: 16
Date: Tue, 2 May 2006 15:42:35 -0700
From: "Randolph-Habecker, Julie"
Subject: [Histonet] RANK, ER and PR in mouse
To:
Message-ID: <21F1955FA284134E981948D6D3FFD0420433EF07@harpo.fhcrc.org>
Content-Type: text/plain; charset="us-ascii"
Histonetters,
Can anyone suggest antibodies to detect Rank, ER, and PR in mouse? If
you have a protocol that you can share, that would be great too!
Thanks,
Julie
Julie Randolph-Habecker, Ph.D.
Staff Scientist - Manager
Experimental Histopathology Shared Resource
Fred Hutchinson Cancer Research Center
1100 Fairview Ave, N. M5-A803
Seattle WA 98109-1024
206-667-6119
jhabecke@fhcrc.org
------------------------------
Message: 17
Date: Tue, 02 May 2006 17:02:20 -0700
From: Yak-Nam Wang
Subject: [Histonet] NADH staining
To:
Message-ID:
Content-Type: text/plain; charset="US-ASCII"
Everyone,
I want to do some NADH staining to determine cell viability in treated
porcine tissue (liver to start with). However, I do not know which reduced
NADH I should be using to make up my solution: alpha or beta. I have seen
papers using both types. Beta-NADH seems to be a lot cheaper but will it
make a difference which one I use?
Thank you for your help
Yak-Nam
--
Research Associate
CIMU
Applied Physics Laboratory
University of Washington
Box 355640
Seattle WA 98195
Tel.: 206-616-6673
------------------------------
Message: 18
Date: Wed, 03 May 2006 01:20:48 -0400
From: John Kiernan
Subject: Re: [Histonet] gallocyanine stain for Nissl
To: Elizabeth Chlipala , Histonet listserver
Message-ID: <44583DB0.9060804@uwo.ca>
Content-Type: text/plain; charset=ISO-8859-1; format=flowed
The procedure in Bancroft & Gamble is OK. You can
shorten the staining time to 2 hours by doing it
in a 60C oven instead of at room temperature. (A
higher temperature may shorten the shelf-life of
the staining solution. Has anyone tested this?)
The correct spelling is gallocyanine (because the
dye is an amine). The CI number is 51030 (CI
Mordant blue 10).
The chemistry of the cationic chromium complex of
the dye and its binding to nucleic acids is well
understood - supported by various studies more
recent than the two cited in B & G. This is a
truly excellent staining method for anyone who's
not in a hurry.
Its big advantages over most other Nissl stains
(such as cresyl violet, toluidine blue and neutral
red) are that there is no overstaining and the
colour resists most subsequently applied chemicals
such as alcohol-water mixtures and acidic
solutions of anionic dyes. Simple basic dyes,
intelligently applied, can provide fast Nissl
staining for small batches of slides. With
chrome-gallocyanine there is no need for speed or
intelligence; that's why I really like the method,
even though the quality of the colour (hue?
saturation?) is a rather bland greyish-blue.
The only shortcoming of the gallocyanine-chrome
alum method is that the staining solution begins
to deteriorate after 1-2 weeks. Because maximum
coloration is confined to the stainable substances
(in the CNS this means nuclear DNA and ribosomal
RNA) it's possible to do mass-production staining
without having to worry about shelf-life. A 400 ml
batch of gallocyanine-chrome alum solution can be
used repeatedly to stain several racks of slides
for a few days. Throw the staining solution out
when you've finished the batch, because if you try
to use it again two weeks later it will be no
good. (Guess how I found that out.)
The intensity of cytoplasmic staining with
gallocyanine-chrome alum has been shown by
scanning microspectrophotometry to be proportional
to the concentration of what we now call rRNA.
Einarson's 1951 paper, which is cited in Bancroft
& Gamble, is a good read, and it shows that there
was molecular biology 13 years before anyone had
seen a ribosome. By the late 1950s it was known
that cells contained more RNA when they were
making more protein. Einarson's method, dating
from 1932, did its bit and is still useful.
John Kiernan
Anatomy, UWO
London, Canada.
--------------------------
Elizabeth Chlipala wrote:
>
> I need to perform a gallocyanin stain for nissel on mouse lumbar spinal
> cord. I have a protocol from bancroft and it says to stain for 18-48
> hours. Can anyone out there give me any pointers on this stain. I also
> need to order the gallocyanin and and the only reagent I see is
> gallocyanine is that the same thing?
>
> 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
>
---
------------------------------
Message: 19
Date: Wed, 3 May 2006 11:50:42 +0100
From: "CRAIG BARLOW"
Subject: [Histonet] Inflamatory markers
To: histonet
Message-ID:
<22482b560605030350v72081d43la6d8f14476551fc5@mail.gmail.com>
Content-Type: text/plain; charset=WINDOWS-1252; format=flowed
Hello All,
I am looking for inflammatory markers in skin that I can perform
quantitative IHC on.
I have a few in mind i.e CD68 just thought I'd throw the question out...
All replies are welcome.
Regards
Craig Barlow
------------------------------
Message: 20
Date: Wed, 3 May 2006 12:14:12 +0100
From: Malam Jacqueline
Subject: [Histonet] Re- processing fatty tissues
To: "Histonet submissions (histonet@lists.utsouthwestern.edu)"
Message-ID:
Content-Type: text/plain
We have used a modified routine method for a few years now and it works fine
with our Shandon Pathcentre processor as long as you keep an eye on the
number of blocks processed and change reagents accordingly, and the blocks
don't exceed recommended size and thickness.
Schedule -
Neutral buffered 10% formalin - 11/2 hrs at room temp (unless you have a
downdraft extractor then use heat)
70% alc, 90% alc then 3 changes of 100% alc (alcohols) - 1 hr all heated to
40 C
1 change of 100% alcohol/xylene mixed ratio of 50:50 - 1 hr heated to 40 C
3 changes of xylene - 1 hr 15 mins all heated to 40 C
4 changes of wax - 1 hr 15 mins all heated to 60 C
Jacqui Malam
Lancaster
UK
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------------------------------
Message: 21
Date: Wed, 3 May 2006 08:12:12 -0500
From: "McGovern, Kevin"
Subject: [Histonet] The end of an era
To: histonet@lists.utsouthwestern.edu
Message-ID:
Content-Type: text/plain; charset=iso-8859-1
Kindly remove me from your mailing list. Thank you.
------------------------------
Message: 22
Date: Wed, 3 May 2006 10:47:40 -0400
From: "Martha Ward"
Subject: [Histonet] Mycobacterium bovis (BCG) antibody
To:
Message-ID:
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My dermatopathologist asked me to look into this antibody to add to our
menu to ccreen for microorganisms. I would appreciate any advice
concerning vendors, conditions, etc. Thanks in advance for your help.
Martha Ward
Molecular Diagnostics Lab
Wake Forest University Baptist Medical Center
------------------------------
Message: 23
Date: Wed, 3 May 2006 11:10:18 -0400
From: Kathie A Berghorn
Subject: [Histonet] Glycerol cryoprotected tissue and cryostat
sectioning
To: histonet@lists.utsouthwestern.edu
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Dear Histo-netters,
I need your help. We have precious tissue that
has been processed for beta-galactosidase. This
was the first time we did this
immunohistochemistry. The protocol said to clear
in glycerol which we did and the tissue is in 80%
glycerol. I have spent the morning trying to cut
this tissue on a cryostat without success. When
I put the mouse placenta in OCT using isopentane
and liquid nitrogen, the OCT freezes but the
tissue does not. Cutting it on the cryostat is
then not possible. When I used to do thick
sections we sunk tissue in sucrose and then froze
it with dry ice prior to cutting it on a freezing
sliding microtome. Would the dry ice freeze the
placenta in the cryostat so I could cut it? Any
other suggestions? Of course I am on a grant
deadline and am stressing.
I would appreciate any suggestions that would
yield 8-10 µm sections so I can see if the beta
galactosidase worked in the area of the placenta
we study.
Thank you very much,
Kathie
------------------------------
Message: 24
Date: Wed, 3 May 2006 17:46:18 +0200 (CEST)
From: Guillermo Palao
Subject: RE: [Histonet] IHC on frozens
To: Histonet
Message-ID: <20060503154618.25177.qmail@web26211.mail.ukl.yahoo.com>
Content-Type: text/plain; charset=iso-8859-1
Hi Sarka,
I have recently had similar problems with parafin vs frozen sections, and these are the conclusions I have come to:
- We routinely store at -20 ºC fridge the frozen sections already fixed for 5 min in acetone at -20 ºC, I do not know if this is the best way to do it, and I would appreciate any more informed input in this sense.
- For quenching endogenous peroxidase you may use H2O2 0.3% (instead of 3%) for 5 min RT (instead of 30 min).
- In my (little) experience, Ab concentrations do not matter that much if the antibody is good. For convenience purposes I stain frozen sections for 1 hour RT as compared with O/N at 4 ºC for FFPE sections.
- We noticed similar (if not worse) background problems due to endogenous mouse IgG in frozen sections, although I did not do a side by side comparison.
I found a really good improvement in terms of background after blocking endogenous biotin with Vector Avidin/Biotin blocking kit. Try staining your slides only with ABC reagent (no Abs at all), and if you see color developing (as was my case) then you definitely need to block endogenous biotin.
Good luck,
Guillermo
Sarka Lhotak escribió:
Hello Netters,
I am trying to get some antibodies work on mouse tissue (see my
previous posting). Since I have not been successful on paraffin
sections I now turned to frozens where I have no experience. I am
getting the same staining (or background of course) on positive and
negative slides (antibody omitted). Even antibodies that I know work
very well on paraffin (my positive control), give me no specific
staining and a background mainly on connective tissue. I've searched
for a proper fixing, drying, dipping in water etc. procedures prior to
staining. I found it very confusing, people swear by different, even
opposing protocols.
So I have a few questions:
What is the best way to prepare/store your sections prior to staining?
Is quenching peroxidase and blocking the same as on paraffin?
Generally, are the primary and secondary antibody concentrations the
same as on paraffin?
Is the MOM problem worse on frozens than on paraffin?
All suggestions are welcome.
Desperately,
Sarka Lhotak
McMaster University
Hamilton, Ontario
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Message: 25
Date: Wed, 03 May 2006 11:49:24 -0400
From: Pamela Marcum
Subject: [Histonet] Save These Dates For The Pennsylvania State
Histology Meeting
To: histonet@lists.utsouthwestern.edu
Message-ID: <6.1.1.1.2.20060503113716.01a00340@mail.vet.upenn.edu>
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Good Morning to All,
We are having the PSH fall meeting in Pittsburgh this year with a special
seminars series on Thursday, October 26th in the afternoon for Histology
Supervisors and Managers to start the meeting. We will offer a CPT Coding
Seminar, a CAP Inspection Seminar and Smart Shopping for Histology For
Contracts Negotiation and Purchasing.
Friday, October 27th we will have a full schedule of speakers for general
and special histology/cytology topics be announced later.
Saturday October 28th we will offer HT Readiness for ASCP Testing (All
Day) and QIHC Readiness Testing (All Day) along with special speakers from
the forensics CSI area throughout the day. We will have topics for routine
and research histology all day.
Join us October 26 to 28, 2006 in Pittsburgh PA.
We are contacting everyone we can in nine states to join us. We are still
early enough in the process to accept suggestions for topics and speakers
for 3 time slots. If you have a suggestion please let me know and we will
consider it.
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
------------------------------
Message: 26
Date: Wed, 3 May 2006 09:49:43 -0600
From: "Elizabeth Chlipala"
Subject: RE: [Histonet] Mycobacterium bovis (BCG) antibody
To: "'Martha Ward'" ,
Message-ID: <001901c66ec9$32794cd0$0300a8c0@Chlipala>
Content-Type: text/plain; charset="US-ASCII"
Martha
We haved used this antibody in research quite a bit on human, mouse and
guinea pig specimens of tuberculosis. We used the antibody from Dako.
It works quite well on FFPE tissue, but there is one thing that you need
to be aware of and that it is not entirely specific to MTB it will also
stain other gram positive bacillia. I believe that is stated on the
spec sheet.
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 Martha
Ward
Sent: Wednesday, May 03, 2006 8:48 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Mycobacterium bovis (BCG) antibody
My dermatopathologist asked me to look into this antibody to add to our
menu to ccreen for microorganisms. I would appreciate any advice
concerning vendors, conditions, etc. Thanks in advance for your help.
Martha Ward
Molecular Diagnostics Lab
Wake Forest University Baptist Medical Center
_______________________________________________
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http://lists.utsouthwestern.edu/mailman/listinfo/histonet
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End of Histonet Digest, Vol 30, Issue 3
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