Protein Fractionation and Identification

The membrane proteins isolated using the colloidal silica method described above are especially difficult to solubilize, and difficult to separate by 2D gel electrophoresis. Consequently the membranes were solubilized in Laemmli buffer with sonication in a 60 °C water bath, and separated by 1D S DS gel electrophoresis. This ID gel was cut into 1 mm bands, each of which was subjected to trypsin digestion. The peptides were recovered, separated and analyzed by LC/ESl MS/ MS. This strategy is illustrated in Eigure 13.5. More than 3200 peptides were 

There are some important differences between the protein inventories developed for the MCF-7 plasma membrane using the shot-gun strategy, and for the MCF-7 nucleus using 2D gel arrays. The shot-gun strategy identified many more proteins. 

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