TCA Precipitation Problems

Chris Trotta trotta at seqaxp.caltech.edu
Fri Aug 18 19:08:15 EST 1995


Hi,

I have been working with small quantities of a nuclear membrane 
protein complex.  In order to get enough to analyze, I must TCA
precipitate.  My problem is after spinning down the protein and 
acetone washing the pellet, I am unable to resuspend the pellet.
I have tried resuspending in dH20, buffer containing Triton X-100
and tris buffer.  No matter what I do to the resuspension, ie heat
to 65 for a few minutes, add SDS loading dye and heating to 90 for
10 minutes, the pellet does not go entirely into solution.  I have
loaded the pellet in the well and it contains a considerable amount
of protein, but this results in an unacceptable smear in the lane.

Does anybody with experience in membrane proteins or protein methods
have any suggestions for getting around this problem???

Thank you,

Chris Trotta
trotta at seqaxp.caltech.edu



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