[Histonet] which one first - antigen retrieval or endo peroxidase block?

[Anne C Lewin]

It doesn't really matter which one comes first - when I am using 
phospho-antibodies the IHC works better when the peroxidase block is 
done _before_ antigen retrieval.  I have done the peroxidase block after 
antigen retrieval but before primary antibody, and also sometimes have 
waited until just before/after  the biotinylated secondary antibody.  
The bottom line is that the peroxidase block needs to be done at some 
point before using the horseradish peroxidase (HRP complex), so that any 
endogenous peroxidase can be quenched and not give you a lot of 
background (especially with any RBC in the tissue).

Rene J Buesa wrote:

>Antigen retrieval always comes first.
>René J.
>
>--- On Thu, 9/25/08, Monfils, Paul <PMonfils <@t> Lifespan.org> wrote:
>
>From: Monfils, Paul <PMonfils <@t> Lifespan.org>
>Subject: [Histonet] which one first - antigen retrieval or endo peroxidase block?
>To: histonet <@t> lists.utsouthwestern.edu
>Date: Thursday, September 25, 2008, 2:26 PM
>
>When doing immunoperoxidase staining on a tissue that requires antigen
>retrieval, which do you do first - the antigen retrieval or the endogenous
>peroxidase blocking?  Or does it matter?
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