immunopptn

R. Woodward rw200 at cus.cam.ac.uk
Thu May 18 08:00:45 EST 1995


Dear all
I am having problems with immunoprecipitation of invitro translated protein products from the Promega TNT coupled system. I get product of the correct size as judged from SDS-PAGE fluorography but I cannot seem to immunoprecipitate the product.  My protein is an ion channel subunit and I have constructed a FLAG tag epitope at the N-terminus, I have varified the cDNA squence and know that the FLAG is expressed, the antiserum has also been tested on western blots using another FLAG tagged protein expressed i
n mammalian cells. I have tried lots of different conditions but even the most mild will not precipitae protein.  Do you think that I need to include pancreatic microsomes in my invitro translation?  Would this have any effect on the anti FLAG recognition?? Any suggestions or advice would be appreciated.
		Thanks Robert (Email rw200 at cus.cam.ac.uk)



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