I am working with primary renal proximal tubule cells and intend to isolate membrane proteins such as sodium bicarbonate co-transporter. What would be the best method compatible with subsequent Western blots?
Many thanks!
hi
RIPA is ok .
good luck
http://www.abcam.com/protocols/sample-preparation-for-western-blot
The methods involved can differ depending on whether you want to identify the integral membrane proteins or peripheral membrane proteins .A cotransporter is likely to be an integral membrane protein.
Integral membrane proteins can be isolated using non ionic detergents like Triton X 114 etc.For peripheral proteins one can use sodium carbonate extraction.
Hi Leta, if you are just looking for a band on a western blot, then you can solubilize the cells in SDS sample buffer, use a small bore needle syringe to shear the DNA by sucking up/expelling the cell lysate through the needle syringe around 10-20 times or until the sample is not viscous. Then Bob's your uncle.
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