I wanna isolate plasma membrane proteins from our cultured cells and send them for mass spectrometry assay for protein quantification. I'm quite naive about protein isolation and mass spectrometry techniques. So, I wanna know, is there any reliable published plasma membrane proteins isolation protocol? And how to perform a mass spectrometry for protein quantification?
I used to purify plasma membranes from CHO cells by sonicating the cells, pelleting all the membranes in an ultracentrifuge, resuspending the whole membranes pellet, then running the extract on a sucrose step gradient. The plasma membranes, being the lightest membranes, formed a band at 16%/31% sucrose interface, whereas the other subcellular membranes went through the 31% sucrose layer.
The more detailed procedure can be found here:
Article ATPase activity of purified and reconstituted P-glycoprotein...
ab65400-Plasma Membrane Protein Extraction Kit
OR
Catalog number 89842-Mem-PER™ Plus Membrane Protein Extraction Kit
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