To enrich for the membrane protein of interest, you can subject the total cell extract to a series of centrifugation steps following physical cell breakage (such as by sonication, rather than detergent lysis). First, a low-speed spin pellets nuclei and unbroken cells. Second, ultracentrifugation of the supernatant pellets all the non-nuclear membranes. Third, sucrose density gradient ultracentrifugation allows the organellar membranes to be separated into fractions because they band at different sucrose concentrations (the plasma membrane is the lightest fraction). If you know which organelle the protein is found in, you can choose the correct fraction. By this method, you will have greatly enriched for the protein you want, and will have an easier time identifying it. I would suggest stating with a lot more cells, say 108.

To enrich for the membrane protein of interest, you can subject the total cell extract to a series of centrifugation steps following physical cell breakage (such as by sonication, rather than detergent lysis). First, a low-speed spin pellets nuclei and unbroken cells. Second, ultracentrifugation of the supernatant pellets all the non-nuclear membranes. Third, sucrose density gradient ultracentrifugation allows the organellar membranes to be separated into fractions because they band at different sucrose concentrations (the plasma membrane is the lightest fraction). If you know which organelle the protein is found in, you can choose the correct fraction. By this method, you will have greatly enriched for the protein you want, and will have an easier time identifying it. I would suggest stating with a lot more cells, say 108.

Thank You very much Shapiro!