Re: [Histonet] double staining for nerve fibers and endothelial cells on skin samples


Au Contraire.  The CD31 (MEC 13.3 rat anti-mouse), staining is much, 
much better the thicker your sections are.  Check out the publications 
from D.M. McDonald from UCSF, he has the absolute best microvessel 
stainings ever published.  If you read the fine print of some of his 
more recent publications you will find two very important clues: 1) 
thick sections 40 to 100 um; 2) Long incubations at room temperature.  
It seems to me you are in luck!  Go ahead with your thick sections, you 
will see blood vessel histochem like you've never seen before.  It 
works like a charm.
Let me know how it goes,
On May 10, 2004, at 11:45 AM, Cao, Xudong wrote:

> Dear all:
> I am trying to determine the time and spacial sequence of nerve 
> innervation and angiogenesis of a grafted skin equivalent using 
> histo-chemical-immunostaining. The primary antibodies that I will be 
> using for this study are PGP9.5 for nerve fiber and CD-31 (i.e. 
> PECAM-1) for vascularization, and I will be using two different but 
> compatible peroxidase substrate to visualize the signals . My 
> questions: 1) is there anything that I need to pay special attention 
> to for double staining in general? 2) the staining for nerve fiber 
> normally calls for a thick section (40 to 50 um) whereas that for 
> endothelial cells calls for a relatively thin section (6-8 um). How 
> can I accommodate this difference in thickness, any suggestions? 
> thanks for the help.
> Xudong Cao, Ph.D.
> Post-doctoral fellow
> Brown University
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