Re: Help How to get rid of background

From:Barbara Wright <bwright@gene.COM>


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Jian

On all of my frozen sections using the ABC method I block the tissues
for endogenous peroxidase with a glucose oxidase method and also block
endogenous biotin with an Avidin Biotin Blocking Kit (Vector). Is your
secondary antibody a goat anti-rabbit IgG?  If not - block the
endogenous immunoglobulins with the serum of the host species of the
secondary.
I hope that helps.
Barb Wright:)

jian ma wrote:

>
>
> Hi, all:
>
> I got lots of background in oral epithelium using ABC method. I
> stained frozen section with a rabbit poly-clonal anti-human antibody.
> The antibody was not commercial now. I have applied high dilution 1 to
> 20 goat serum for1 hour for blocking. The color was developed in DAB
> for 30 second to 1 minute, The background staining in the epithelium
> still can be seen. It was said that rabbit serum, but not mice and
> goat, contains environmental antibodies to human epithelium or Fossman
> antigen reaction. Is it impossible to get rid of background?
>
> Any suggestions will be very thankful.
>
> Jian Ma
>
>
> -----------------------------------------------------------------------
> Get Your Private, Free E-mail from MSN Hotmail at
> http://www.hotmail.com.

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Jian
<p>On all of my frozen sections using the ABC method I block the tissues
for endogenous peroxidase with a glucose oxidase method and also block
endogenous biotin with an Avidin Biotin Blocking Kit (Vector). Is your
secondary antibody a goat anti-rabbit IgG?  If not - block the endogenous
immunoglobulins with the serum of the host species of the secondary.
<br>I hope that helps.
<br>Barb Wright:)
<p>jian ma wrote:
<blockquote TYPE=CITE> 
<p>Hi, all:
<p>I got lots of background in oral epithelium using ABC method. I stained
frozen section with a rabbit poly-clonal anti-human antibody. The antibody
was not commercial now. I have applied high dilution 1 to 20 goat serum
for1 hour for blocking. The color was developed in DAB for 30 second to
1 minute, The background staining in the epithelium still can be seen.
It was said that rabbit serum, but not mice and goat, contains environmental
antibodies to human epithelium or Fossman antigen reaction. Is it impossible
to get rid of background?
<p>Any suggestions will be very thankful.
<p>Jian Ma
<p><br>
<hr>Get Your Private, Free E-mail from MSN Hotmail at <a href="http://www.hotmail.com">http://www.hotmail.com</a>.</blockquote>
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