[Histonet] Re: Histonet Digest, Vol 1, Issue 187

Joanne Mauger MAUGER <@t> email.chop.edu
Mon Dec 29 09:40:46 CST 2003

What animal is your secondary made in? Try blocking in the normal serum of that animal for 10 minutes before applying primary ab.

>>> "Amos Brooks" <amosbrooks <@t> earthlink.net> 12/27/03 09:42AM >>>
         That really sounds like endogenous biotin. Try either a biotin 
block or a non biotin detection system. You can get the biotin block from 
several companies (I know DAKO carries it) there's also a home made 
blocking recipe using milk. Search the archives if you want to try the milk 
thing. It's been discussed here before.
         The non-biotin detection systems are also a good choice. You could 
use either alkaline phosphotase or one of the new systems that use a 
dextran polymer like Envision from DAKO (there are other companies that 
carry it but it's Saturday and my mind is currently on cartoons and cereal 
:-) )
Good luck,
Amos Brooks

At 01:00 PM 12/26/03, you wrote:
>I'm doing IHC on rat skeletal muscle. I got a heavy background stain on 
>all sections and control section without 1st AB. But there is no stain on 
>the control section without 2nd Ab.
>It looks like the 2nd Ab will react with the rat tissue. My first Ab is 
>derived from Mouse, and second Ab is anti-mouse.
>What could I do to reduce the background stain?

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