[Histonet] Mouse GranzymeB

Tue Jan 28 13:10:31 CST 2014

Amos u might have to get an anti rabbit link that has been mouse absorbed.
Dako's anti rab envision is made in a goat but it is not mouse absorbed.
Southern Biotec or Jackson Labs are really good places to get absorbed
links.  Still no reliable cd4 and cd8 for ffpe mouse tissue as far as I
know, I have tried many that claim they work but have not consistently
worked in my hands, we still do that with frozen mouse tissue not aldehyde
fixed.  Are u doing HIER with the EDTA ph9?  In my experience especially
with the higher ph u can over retrieve and cause undesirable nuclear
staining.

Patsy Ruegg, HT(ASCP)QIHC
Ruegg IHC Consulting
40864 E. Arkansas Ave
Bennett, CO 80102
H 303-644-4538
C 720-281-5406
pruegghm <@t> hotmail.com 
rueggihcconsultingpr <@t> outlook.com

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-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Amos Brooks
Sent: Friday, January 24, 2014 10:09 AM
To: histonet <@t> lists.utsouthwestern.edu; ihcrg Group, (E-mail)
Subject: [Histonet] Mouse GranzymeB

Ok Musketeers,
     I am trying to detect cytotoxic T-Cells in formalin fixed paraffin
embedded mouse liver. GranzymeB should do the trick. I have a rabbit anti
mouse (and human and rat) GranzymeB from abCam (ab53097). The spleen that I
ran as a control with it works fine. Nice T-cells and no signifigant
background staining. The liver on the others hand has some lymphocyte
staining, presumably cytotoxic T-cells as expected,  but it looks like every
hepatocyte nuclei are also picking it up.  Grrr,  right?
     If this antibody had been raised in mouse,  I wouldn't be surprised.
Also,  if the spleen looked similar, I wouldn't be surprised. Do any of you
have any ideas ab pi ut what this might be? Alternatively, does anyone have
any ideas about another antibody that would work here?  CD4 or CD8 would be
great here,  but its FFPE mouse,  so sadly that's out (more grumbling).
    For the completionists out there,  I used EDTA pH9,  peroxide block and
detected the antibody (diluted 1:800) with Envision (rabbit) and DAB from
Dako.

Any suggestions would be vastly appreciated,

Happy Friday,
Amos
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