[Histonet] blocking endogenous activities

[Jan Shivers]

You need to do it sometime before the HRP-labeled conjugate is put on.  If
you put it on after the HRP conjugate, you'll block the HRP substrate and
won't see where the primary antibody is binding.  I usually do mine before
antigen retrieval, but have done it after the primary antibody step on
frozen sections and cell smears.

Jan Shivers
Senior Scientist
IHC/Histology/EM
Veterinary Diagnostic Lab
University of Minnesota
St. Paul, MN

On Thu, Mar 1, 2012 at 5:55 PM, Bea DeBrosse-Serra <
BDeBrosse-Serra <@t> isisph.com> wrote:

> The majority of the time we do it after retrieval. I believe you can do
> the hydrogen peroxide block at any point, as long as you do it before the
> DAB.
>
> Beatrice DeBrosse-Serra HT(ASCP)QIHC
> Isis Pharmaceuticals
> Antisense Drug Discovery
> 2588 Gazelle Ct.
> Carlsbad, CA 92010
> 760-603-2371
>
>
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:
> histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Tuyen Nguyen
> Sent: Thursday, March 01, 2012 3:49 PM
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] blocking endogenous activities
>
> In my lab, in IHC protocols for most antibodies, hydrogen peroxide is
> applied before antigen heat retrieval. However, I wonder there is any
> different result in IHC stain if hydrogen peroxide for blocking endogenous
> activities is applied after antigen heat retrieval by using steamer or
> pressure cooker?
> Thank you,
> Mai Nguyen Truong
> Research Associate II
> Los Angeles, California
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