(by tullum from cfl.rr.com)
Fri Feb 16 10:02:40 EST 2007
"atahualpa" <botulina from poczta.onet.pl> wrote in message
news:Xns97C8BB8867C05sralwaspiesgazetapl from 126.96.36.199...
> I have a following problem with immunoprecipitation:
> When I develope western of the cell extract with the antibodies for the
> protein I am dealing with I see the protein that I expect and two, much
> thicker, bands higher. I want to immunoprecipitate them and go for MALDI.
> have run immunoprecipitation of the cell extract with the same antibodies
> and I run gel for coomasie staining. I can see on it very nice band of my
> protein and nothing more beside background. It seems that IP works since I
> can catch my protein, but where are the other two guys? Why they are not
> caught in IP although very nicely detected on western with same
> The only reason that comes to my mind is that western is developed in
> conditions different from IP (milk vs lysis buffer). Could it be that
> buffer (contains 1% NP40) affects binding conditions of those two not
> affecting binding of the "original" protein?
> I would appreciate all suggestions.
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