Separation of Membrane Vesicles and Cytosol from Yeast, Cultured Cells, and Bacteria in a Small Volume Self-Generated Gradient in a Fixed-Angle Rotor

Abstract

There are many situations when it is necessary to separate rapidly and efficiently a cytosolic and a membrane vesicle fraction from yeast, cultured cells, or from bacteria. This Protocol Article describes the flotation of the vesicles through a self-generated gradient from a dense sample zone using the low-viscosity medium iodixanol. As the sample is exposed to the g_max the tendency of the proteins to sediment overcomes any diffusion in the opposite direction and are therefore completely separated from the vesicles.