elution of co-IP from protein G beads

Pow Joshi via methods%40net.bio.net (by pow.joshi from gmail.com)
Thu May 29 12:15:00 EST 2008


On 28/05/2008, AK <akhan357 from sbcglobal.net> wrote:
>
> Dear experts, I am doing a co-immunoprecipitation experiment. I pull down
> protein X with a mAb and detect co IP of protein Y with pAb. protein Y is
> heat sensitive and if I heat (~70C) the sample to elute immune complex, I
> can not detect protein Y. without heating, protein-G beads do not let the
> IgG chains loose, at least. Are there other ways to elute immune complex
> without heating? of course low pH is one obvious strategy that I have not
> tried yet. thank you in advance.



could you perhaps use high salt say ~500mM for elution? You would only need
to break the mAb-proeinY binding.....?

best
Pow

AK
>
>
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