[Histonet] Re: Histonet Digest, Vol 10, Issue 16- New question


Hello Everyone-

We are starting up an IHC department in a Vet. Lab and was wondering if animal tonsils or lymph nodes work for T and B cell markers, or what is working in your animal lab for CD3, CD45, and CD79a?  Thanks in advance.

-Beth Delescavage, BS, HTL
 Phoenix Central Labs
 Everett, WA

----- Original Message -----
From: histonet-request@lists.utsouthwestern.edu
Sent: Saturday, September 11, 2004 10:03 AM
To: histonet@lists.utsouthwestern.edu
Subject: Histonet Digest, Vol 10, Issue 16

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Today's Topics:

   1. RE: Question regarding 10% Neutral buffered formalin/Immuno
      (Barry R Rittman)
   2. (no subject) (Giorgia Setti)
   3. Nemesis stainer (Olszewski, Dawn)
   4. Re: Tissue transport from processor to embedding station
      (Amy Porter)
   5. Work load for research labs doing GMA (Macke, Gail)
   6. IHC on Frozen Sections (Allen, Rhonda)
   7. GFP in a fixed brain tissue (Ajai Vyas)
   8. Re: Leica 1800 cryostat manual
   9. Off Topic - question about class action suit against your
      "employer" (Victoria Baker)


Message: 1
Date: Fri, 10 Sep 2004 12:16:58 -0500
From: "Barry R Rittman" 
Subject: RE: [Histonet] Question regarding 10% Neutral buffered
To: "histonet" 
Content-Type: text/plain; charset="us-ascii"

The original idea of adding formalin or glutaraldehyde to the EDTA was
to prevent maceration during the long exposure to EDTA solutions. In my
experience these aldehydes significantly decrease the rate of EDTA
demineralization. This may well be due to increased cross linking,
especially if the original fixation was of relatively short duration.
Have no evidence or references for this. My advice would be to use EDTA
pH7.2 to 7.4 to eliminate this as a potential problem.

-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu
[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of
Dunn-Jena, Patsy A
Sent: Friday, September 10, 2004 9:49 AM
To: Histonet
Subject: [Histonet] Question regarding 10% Neutral buffered

I have a graduate student who has come to me with a question I cannot
answer as I have limited Immunohistochemical experience.  One of her
advisers wants her to go to using an EDTA decalcification recipe which
contains the 10% NBF in it which is a recipe we used on all of his
previous work.  She asked me about the NBF crosslinking her immuno sites
as she has been using an EDTA decalcification only recipe.  Some input
here would be nice so I can better explain it to her.

Patsy A. Dunn-Jena, RVT, LAT, HT (ASCP)
Mineralized Tissue and Histology Research Laboratory
Indiana University School of Dentistry
1121 W. Michigan Street, Room 238
Indianapolis, IN 46202

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Message: 2
Date: Fri, 10 Sep 2004 18:23:54 +0100
From: Giorgia Setti 
Subject: [Histonet] (no subject)
To: histonet@pathology.swmed.edu
Message-ID: <1094837034.4141e32a9fa7a@impmail.kcl.ac.uk>
Content-Type: text/plain; charset=ISO-8859-1

Giorgia Setti

hello histonetters!!!!
i am desperate because I can't find a good antibody for MCP-1 staining on rat
frozen sections;I tried the MCP-1 antibody of Abcam but it didn't work;do you
know any other antibody tested on frozen section??
thank you Giorgia

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Message: 3
Date: Fri, 10 Sep 2004 13:29:18 -0400
From: "Olszewski, Dawn" 
Subject: [Histonet] Nemesis stainer
To: "'histonet@lists.utsouthwestern.edu'"

Message-ID: <825F2CDD64ACF34D85E5E5CCC9EA01320557EA2B@email.sgmc.org>
Content-Type: text/plain

Does anyone have any positive or negative comments about the Nemesis IHC
stainer?  Any info would be appreciated.  Thanks in advance.

Dawn Olszewski HTL(ASCP)QIHC
South Georgia Medical Center


Message: 4
Date: Fri, 10 Sep 2004 10:45:54 -0700 (PDT)
From: Amy Porter 
Subject: Re: [Histonet] Tissue transport from processor to embedding
To: Maria Christensen ,
Message-ID: <20040910174554.28637.qmail@web40901.mail.yahoo.com>
Content-Type: text/plain; charset=us-ascii

Maria - reply to me (off list) if you want and supply me with your fax number and i can fax you pictures of what we have here.  It is a Thermal Console we have model #4585 from Miles/Sakura.  We also walk several feet to our embedding station and always have just placed our cassettes into the reservoir then let them sit for a few minutes to rewarm.  They usually do not get that cold in that short of a time span.

Maria Christensen  wrote:
Thanks to all who replied. To answer some of your questions, our  
processor and embedding center are in separate rooms now, but not far  
from each other (10-12 steps). Our concern was keeping the tissue  
warm so that we could embed quickly when needed without having to  
warm the tissue up. We thought of just transferring in a beaker of  
melted paraffin, but sometimes we can't get to all the cassettes  
right away, so the wax would begin to solidify. Unfortunately we  
don't have an oven in the lab where the embedding equipment is.

Our Tissue Tek II embedding station has a heated compartment for the  
molds, but no compartment for the cassettes. The manual mentions an  
infiltration process carrier, and a vacuum infiltrator, but these  
accessories did not come with our reconditioned equipment. I would  
like to know more about the holding reservoir that Amy Porter  
mentioned - part no., where purchased, etc. because if the basket  
from the VIP fits in it, that would be ideal.

Again, thanks for your ideas- I'm a firm believer in not reinventing the wheel!


Maria A. Christensen
Technical Associate
Medical Microbiology & Immunology
Creighton University School of Medicine
2500 California Plaza
Omaha, NE 68178-0404

phone (402) 280-2678
FAX (402) 280-1875
email mariac@creighton.edu
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Amy S.Porter, HT(ASCP)  
Michigan State University  
Department of Physiology  
Division of Human Pathology  
College of Human Medicine  

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Message: 5
Date: Fri, 10 Sep 2004 14:06:03 -0400
From: "Macke, Gail" 
Subject: [Histonet] Work load for research labs doing GMA
To: "'histonet@lists.utsouthwestern.edu'"

Content-Type: text/plain; charset="iso-8859-1"

Dear Histonetters,
      Could some one tell me what is considered the average work load for
cutting GMA blocks.  How many Blocks per year embedded? How many slides cut
per year and stained per year? We use Technovit 7100 for processing and
embedding. We are a Core Research Lab that also does paraffin and frozen
processing and cutting.  
       Most of the information I have found relates to Clinical Labs doing
paraffin and frozen work. I haven't found much related to Research Labs.
       Any information would be helpful.
              Thank you for taking the time to read and reply to my
                Gail Macke, HTL
                Shriners Hospital for Children: Shriners Burns Hospital-
                Cincinnati, Ohio, USA
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Message: 6
Date: Fri, 10 Sep 2004 12:30:45 -0700
From: "Allen, Rhonda" 
Subject: [Histonet] IHC on Frozen Sections
To: "'histonet@lists.utsouthwestern.edu'"

Message-ID: <707BC118BC44D3118E3B0000F81F1E7605A393DA@VHAKANEXC1>
Content-Type: text/plain


    I am trying to do the T & B cells on frozen section tonsil for my IHC

    due at the end of this month.  I have tried multiple procedures, and

    not yet found one that makes me happy, or looks like the one in the

    at the IHC home page.

    If someone could e-mail me their procedures (mostly from cutting to

    application) it would be greatly appreciated.

    By the way, my cryostat will not cut thinner than 4-6 microns.

    Thanks in advance,

    Rhonda Allen

    VA Medical Center


Message: 7
Date: Fri, 10 Sep 2004 18:12:43 -0700
From: Ajai Vyas 
Subject: [Histonet] GFP in a fixed brain tissue
To: histonet@lists.utsouthwestern.edu
Message-ID: <934be6370409101812725ddc95@mail.gmail.com>
Content-Type: text/plain; charset=US-ASCII

   Regards from a new entrants to histonet. I have GFP labelled
parasites sitting in brain of my rats. I need to slice the brain and
locate these parasites. Will GFP still be present if I use fixatives
like formaldehyde or organic solvents like xylene? What fixing
protocols are best for preserving GFP signal?
Thanks in advance,


Message: 8
Date: Fri, 10 Sep 2004 11:20:57 -0500
From: Don.Birgerson@leica-microsystems.com
Subject: Re: [Histonet] Leica 1800 cryostat manual
To: "Amy Self" 
Cc: histonet@lists.utsouthwestern.edu,

Content-Type: text/plain; charset=us-ascii

Hi Amy,
      I be happy to mail you a copy of the "User Manual" if you will give
me your street address.
Best regards,

Don Birgerson
Leica Microsystems
Technical Assistance Center
1-800-248-0123  ext 5918

                      "Amy Self"                                                                                                    
                      Sent by:                              To:                                                   
                      histonet-bounces@lists.utsouth        cc:                                                                                      
                      western.edu                           Subject:  [Histonet] Leica 1800 cryostat manual                                          
                      09/10/2004 09:25 AM                                                                                                            

Does anyone have a manual for the old Leica 1800 cryostat that they could
share with me or don't need anymore?  We       (biomed dept.) has
misplaced ours.   Thanks,  amy

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Message: 9
Date: Sat, 11 Sep 2004 06:53:05 -0700 (PDT)
From: Victoria Baker 
Subject: [Histonet] Off Topic - question about class action suit
against your "employer"
To: HistoNet Server 
Message-ID: <20040911135305.17671.qmail@web50105.mail.yahoo.com>
Content-Type: text/plain; charset=us-ascii

Good morning,

This is a question that I hope very few of us on the
list-server have ever had to, are currently, or will
ever have to deal with/answer personally.   

A week ago yesterday the not-for-profit/NCI (Cancer
Center Support Grant)funded organization I work for
gave us WARN notification that told us we would be
permanently "layed-off" as of Sept. 10 unless an
agreement between our facility and the government was
made. As negotiations were being worked for a merger
with "other" institutions there might still be a way
to work out our difficulties and keep our institution
open.  The specifics of this merger are unknown and we
have had no specific information given to us.  As of
yesterday afternoon we were informed that we would
have to call in on Monday morning to see if we would
be allowed access to the building and work.   

The controversy is this:
1.  According to law we are to be notified at least 60
days in advance of a permanent closing.  This has not
been done.
2.  We lose any severance or unused vacation pay still
owed to us under a "normal" - I use this term loosely
- closing.
3.  Insurance benefits - being able to apply for COBRA
or get clear information as to what we could do to
purchase insurance on our own has been non existent
and our HR person left her position as of Friday as
well.  This is not part of the actual legal
proceedings just concerns of all employees.   
4.  Retirement plans/annuities are a separate issue as
not all employees were involved in these benefits.

We are a very small institute of less than 150 people
in NYS.   

I'm looking to see if anyone else on the server has
had this occur to them and whether or not they did
pursue a legal action to get the benefits due to them.
I'd also like to know if anyone has information or
people/places I could contact about what most have
gotten in terms of litigation. The issues I listed are
the most pertinent, as with many of these closings
other issues revolve around the main ones.   

Many of these people have been here since the late
70's or early 80's.  Yesterday as most of us were
cleaning out offices and trying to organize our labs,
I encountered so many different human reactions and
emotions.  When I left my lab last night it was just
around sundown and shadows were cast over some of my
equipment and benches after I had shut down the lights
(I have a wall of windows that I've always loved to
watch the seasons or the weather)and it was a very sad
moment. Would I ever see it again?  When you are layed
off or a company closes it's a 'done deal' and you can
move on.  When you just seem to get a 'stay of
termination' that leaves you no where closer to
resolution it wears you down.     

If anyone has anything that they could pass on I would
very much appreciate it.   

Thank you very much.

Vikki Baker
Institute for Cancer Prevention
Valhalla, NY  10595

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