Thanks for your reply. We are getting great staining with our CD31's. The
problem is the researcher wants an antibody that is exclusive to
endothelial cells of mouse (ie.....no monocytes, etc.)  and it should not
cross react with human endothelial cells. (it's a xenograft, human on
mouse). Any ideas are appreciated.  Denise Woodward


On Fri, 3 Dec 1999 08:49:17 -0700 Gayle Callis
<uvsgc@msu.oscs.montana.edu> writes:
> Have a grad student who does this more than frequently, now doing it
> with confocal.  Frozen sections, cut at 5 um, cold acetone fixation
> for 10 min, refrig temps, air dry 30 min before fixation, fix then
> air dry for another 30 min.  Staining is done (you will need to do
> a dilution panel), using Dulbeccos PBS, pH 7.4 with 0.2% goat serum,
> block is 5 - 10% goat serum 30 min,  staining with antibody is 30
> min,
> Secondary is goat antiRat F(Ab')2 biotin, 1:250 from Biosource
> (Jackson
> has excellent secondary also), Strepavidin-HRP 1:500 from Biosource
> 20 min,
> then chromogen of choice, we like Vector AEC or Pierce enhanced DAB.
> Go easy on hematoxylin counterstain.  If you can add Tween 20 to
> buffer,
> 0.05% to 0.1% and we do both peroxidase block (Dako) and avidin
> biotin block)
>
>
> Good luck
> Gayle Callis
>

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