[Histonet] background in lymphoid tissue

[Johns, Laura [CNTUS]]

Hi Liz,

Thanks for your response.  I am using normal goat serum in place of the
primary and secondary antibodies.  I dilute my primary in NGS, and my
secondary in diluent.  I haven't tried adding in another protein block or
power block; I really thought the avidin/biotin would take care of it, but
it didn't, and I haven't had a chance to try anything else yet.  Any
suggestions are welcome!

Laura

-----Original Message-----
From: Liz Beitman [mailto:LBeitman <@t> cellmarque.com]
Sent: Friday, February 10, 2006 1:18 PM
To: Johns, Laura [CNTUS]
Subject: RE: [Histonet] background in lymphoid tissue


Hi Laura,

Just curious here, what might you be using in place of applying antibody
to the negative?  Might you be using a mouse or rabbit negative serum?
Sometimes these serums are more sensitive than if you just used diluent.
There are other types of blockers as well such as protein blocker that
blocks for any endogenous proteins that can be inherent to certain
tissues.  Have you tried a different blocker?  I hope you don't mind my
response and if there is anything I can do to help, please feel free to
give me a call anytime.

Sincerely,

Liz Beitman
Cell Marque
1-800-665-7284 x 19 

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Johns,
Laura [CNTUS]
Sent: Friday, February 10, 2006 12:04 PM
To: 'histonet <@t> lists.utsouthwestern.edu'
Subject: [Histonet] background in lymphoid tissue

Hi All,

I am having a problem with background staining in lymphoid tissue.  I
have tried increasing the time of my peroxide block, and adding in an
avidin/biotin blocking step, but still there is some slight staining in
my negative controls (no primary or secondary antibodies).  I have
noticed this in spleen and in other organs where there is an area of
inflammation, and with several different antibody combinations.  Has
anyone experienced this problem?  Any suggestions for getting rid of it?

Thanks!
Laura

Laura M. Johns, Ph.D.
Centocor, Inc.
145 King of Prussia Road (Mail Stop # R-4-2) Radnor, PA 19087
Phone:  610-240-8284 ....  Fax:  610-240-8150....  Email:
ljohns53 <@t> cntus.jnj.com

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