12CA5 MAb immunoprecipitation

Jon Backer backer at aecom.yu.edu
Mon Sep 4 11:35:16 EST 1995


In article <Pine.3.89.9508301945.A6736-0100000 at eukaryota>, massonjy at rsvs.ulaval.ca (Jean-Yves Masson) says:
>
>
>Dear Netters,
>
>        We are trying to immunoprecipitate (from total yeast lysate) an 
>epitope tagged protein.

In proteins where the HA tag works, we have not needed anything special for IPs.
However, the thing to note is that results with HA can vary quite a bit protein
to protein. We have had some in which it works for both IP and blot, and some
where it doesn't work at all. If your HA-tagged protein works for blot but not
IP, you might try IPing under denaturing conditions - denature in boiling 1% SDS/10 mM Tris 7.4, then dilute
down to 0.1%SDS/1% Triton X-100/150 mM NaCl/1mM EDTA plus inhibitors and other goodies as needed.
Also, some people use mutliple HA tags in tandem for improved precipitability.

Good Luck.

Jon Backer



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