Does anyone have a simple and cheap protocol for isolating the membrane fraction Vs the cytosolic fraction of adherent cultures of mammalian cells?

11 May 2021 1 7K Report

I am looking hopefully for numbers here: type of plate used, volume of lysis buffers used at each step, formulation of lysis buffers etc.

I have found protocols for hypotonic lysis of suspension cultures with separation of membrane fraction by simply sedimenting in the hypotonic lysis buffer, calling the supernatant the cytosolic fraction, and solubilizing the pellet in detergent as the membrane fraction. .

I am trying to scale this to my adherent cultures, but it would be helpful if someone already has.

On the bright side, I will be over-expressing the target proteins. So far the protocols I have found start with 2*10^8 cells, and that will be problematic for me. I hope to start with less cells.

Thanks

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