I always find it's very difficult to adjust similar co-transfected vs single transfected protein expression amount for co-ip several different constructs. (My boss's absolute requirement before actually doing co-IPs) like:
Protein A + vec
Protein B + vec
Protein C + vec
A+B (both upregulated)
A+C (both upregulated, but A has much more upregulated level than A+C and A alone)
So, the level of A will never be the same in all cases (A, A+B, A+C)!
One of my labmates suggests me to simply mix single protein-transfected HEK293 lysate.
Incubate the mixed lysates for 1hr and then add antibody to do Co-ip.
Do you guys consider it is cheating or an acceptable method? (in theory, I think it's acceptable...)
Edited by Jason S LEUNG, 09 January 2011 - 01:14 AM.