We are performing cell fractionation to separate several organelles + cytosol + plasma membrane- We then want to isolate peripheral proteins from plasma membrane.
We have tried increasing pH (up to 13) and ionic strength with KCl, which as I know are the most used methods. However, when checking specific markers for each fraction, we are not able to see our peripheral protein marker (Ras) in the peripheral protein fraction, but only in the membrane fraction. So, our peripheral proteins are not separated from plasma membrane.
Do you know alternative methods or protocols? We are looking preferably for non-denaturing and non-disrupting membrane methods, but any proposal or idea would be appreciated.
Thanks.
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