Co-Immunoprecipitation of nuclear receptor

[Zhicheng Dong]

If you use formaldehyde as crosslinker, you can break the interaction 
by heating. You can compare the sample with and without heating. 
Without heating your complex will be intact. One thing keep in mind 
is once you do crosslinking, it will be really hard to extract the 
protein that you are interested in. You'd better do time course to 
optimize the condition for crosslinking.

Zhicheng

  At 05:24 PM 3/4/2007, Nath wrote:
>Hi everyone,
>
>I'm presently having problem to do my co-immunoprecipitation. I
>immunoprecipitate a nuclear receptor and I'm trying to look at the
>proteins that are associated to that receptor. The problem is that,
>once actif, my receptor become highly link to DNA, so it become
>insoluble and the condition that affectes its DNA-protein interaction
>also distroyes the protein-protein interaction with my protein of
>interest.  I was wondering if I could crosslink my proteins before the
>immunoprecipitation (to have the active form of my receptor) and look
>at it after by SDS-Page ? Does the condition of an SDS-page is harsh
>enough to break all the protein-protein interaction of crosslink
>proteins so that I can look by western blot at my protein of
>interest ?
>
>Thank in advance,
>
>Nath
>
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