Re: Daily Digest

From:Gayle Callis <>

CD31 (PECAM) is available from several sources, Pharmingin, probably
Serotec, eBioscience.  

Zinc fixation is NOT ZINC FORMALIN, often confused because of the way
Pharmingen put it in their catalog.

Zinc TRIS Buffer fixation needs to be tested, not large tissues, I suggest
a fixation time study, up to optimal time.  If tissue is not fixed by this
"fixative" then fixation will continue during paraffin processing.  You
will get decent morphology, but sectioning is a bit difficult as tissues
are drier, crunchier.  A good ice water soak is necessary, be sure to fix
small, thin pieces of tissue and don't overprocess (too long in dehydrants,

You get primary antibody concentrations similar to acetone fixed frozen
sections, NO ANTIGEN RETRIEVAL necessary.

You can use whatever Alk phos chromogen you want, DAKO has a nice one
called Fuchsin +, a bit more sensitive, red, excellent, we use this with
their SA-AP, inhouse dilution. We tried this with Vector BCIP/NBT with good
results on murine T cell marker.  Chromogen is what you prefer, red or blue
color, with or without hematoxylin counterstain. 

Levamisole quenches endogenous alk phos in tissues with exception of gut
alk phos, which is quenched by acetic acid (have done this on mouse gut,
fixed with Zinc TRIS Buffer, was lovely) 

You should access the original articles, Nitta et al, for technic and
antibody dilution ranges, was helpful.  Pharmingen catalog has those

At 04:25 PM 12/8/00 +0200, you wrote:
>I need to visualise CD31 in rat tissue. There appears to be only one
>monoclonal available from Research Diagnostics. It requires zinc fixation
>prior to paraffin embedding and  alk. phos detection.
>Does anyone have an opinion on
>1. Zn fixation
>2.what is the optimal chromogenic substrate for alk phos
>3. Dako mentions an endogenous alk phos inhibitor, levamisole which they
>put in some of their kits
>Nazlia Samodien
>Cardiovascular Research Unit
>Dept. of Cardiothoracic Surgery
>University of Cape Town Medical School
>Anzio Rd, Observatory, 7925, South Africa
>Tel. +27 21 406 6398 (office)
>        +27 21 406 6476 (Secretary)
>Fax + 27 21 448 5935
Gayle Callis
Veterinary Molecular Biology
Montana State University
Bozeman MT 59717-3610
406 994-4705
406 994-4303

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