co-immuno-precipitation

Bernard Murray bpmurray*STUFFER* at socrates.ucsf.edu
Wed Jun 10 18:55:08 EST 1998


In article <357ED08F.6A74E887 at ccc.uab.edu>, xianghuai lu
<xianghuai.lu at ccc.uab.edu> wrote:

> Hi, groups:
>   I am doing an experiments trying to co-IP two proteins and then do
> western blot to check them. One protein is 57 KD (CD5) and the other is
> 100 KD (Ap2). It looks like I can use larger one to bring down smaller
> one but not the other way around. The other way is what we wanted. Does
> anyone have an better idea to do IP or any tricks?
> Any suggestion will be greatly appreciated.
> Lu

Any chance the interaction site on the small protein is the
same as the epitope for your antibody?  This would mean that
the larger one would be blocked from interacting.  If you
are using a polyclonal antibody for immobilising the small
one this should not be a real problem (unless the interacting
site is the major epitope).
     You may be able to test this by seeing if the anti-CD5
antibody blocks co-IP by the anti-Ap2 antibody.
     If you have the option then try a different antibody
or a different method of immobilising the small one.
     Good luck,
          Bernard
-- 
Bernard Murray, PhD
Dept. Cell. Mol. Pharmacol., UCSF, San Francisco, USA



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