[Histonet] Pericyte, smooth muscle cell IHC problems

[Jackie M O'Connor]

HEY!  I've had the same problem with the Santa Cruz CD31!  How utterly 
frustrating!  I had a lot from 11/04 that didn't work, and when I 
requested a replacement this month, that didn't work either!!  I've really 
come to depend on that antibody for microvessel density in my xenografts. 
I haven't been able to find a comparable CD31 to work in FFPE mouse.  The 
BD will work on zinc fixed, tho, without antigen retrieval.   Anyone else 
have any tidbits for FFPE murine tissue?

Jackie O'
Jacqueline M. O'Connor HT(ASCP)
Assistant Scientist
GPRD Cancer Research
Abbott Laboratories, Abbott Park, IL
Jackie.OConnor <@t> abbott.com




"Melissa Gonzalez" <Melissa.Gonzalez <@t> cellgenesys.com>
Sent by: histonet-bounces <@t> lists.utsouthwestern.edu
05/05/2005 12:19 PM

 
        To:     <histonet <@t> lists.utsouthwestern.edu>
        cc: 
        Subject:        [Histonet] Pericyte, smooth muscle cell IHC problems


Dear all, 
I have been trying unsuccessfully to stain for pericytes/ smooth muscle 
cells in sub cutaneous gliomas in a mouse model. (Tissues are either 4% 
PFA perfused or 10% FFPE)
I have tried several products, including:
* NG2 (Chemicon): works nicely in normal mouse organs, but binds to much 
of the tissue in the gliomas, making it impossible to interpret. 
*PDGFRB (Santa Cruz): resulted in a few crazy-appearing vascular areas, 
very weird
* ASMA (DAKO, Sigma, US Bio, Neomarkers, Abcam)
*Desmin (DAKO, Santa Cruz)
--in both monoclonals (using a mouse on mouse kit which did not work) and 
polyclonals, for which I only see staining in the attached skin and a very 
rare area in the tissue. I have tried both immunofluorescence on the 
perfused,  and chromogenic detection on paraffin sections, using Citrate, 
high pH, pepsin, and proteinase K retrievals. 
 
Is it possible that smooth muscle cells would not be well developed around 
blood vessels in a sub cutaneous glioma, and that is the reason I only see 
staining in the attached skin with a rare occurrence in the tumor? (Tumors 
are very vascular, blood vessels are definitely present)
 
Also, what is everyone using for blood vessel markers? We used to use BD 
Pharm. Pecam-1, which doesn't work anymore on anything except frozens, 
Santa Cruz goat Pecam-1 was working great but the most recent lots I 
ordered now appear completely negative using any detection. I have now 
switched to using CD105.
 
Any help would be greatly appreciated as I have tried all the tricks I 
know, and am left completely puzzled. 
 
Thanks so much, 
Melissa
 
 
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