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thank you for the manifold and quick response to my question.
First of all, I performed Western blotting with an anti-phospho p38
antibody (New England BioLabs). They said that this antibody recognized
only the dual phosphorylated form of p38. I get a very nice
phosphorylation kinetik with this method.
Next, I take the same lysates and performed immunoprecipitations with
the p38 MAP kinase assay kit from New England Biolabs. The anti-phospho
p38 antibody was immobilized by crosslinking to crosslinked agarose
hydrazide beads. ATF-2 was used as substrate in the non-radioactive
kinase-assay, and detections were performed with anti-phospho ATF-2
NO phosphorylated ATF-2 was detectable!!! So I suggest that the
phosphorylated kinase is not active.
Has anyone used this p38 MAP kinase-assay kit, too???
- Re: p38
- From: DR JULIA GEE <Gee@cardiff.ac.uk>