Comparison of Phase Partition with Alternative Methods for Isolation of Subcellular Structures

Abstract

Preparative free-flow electrophoresis alone or in combination with phase partition provides opportunities for decisive separations of membranous cell components and organelles based on surface charge not easily duplicated by classical differential or density centrifugation techniques. Applications include subfractionation of Golgi apparatus into component cisternae, isolation of plasma membrane and tonoplast vesicles from plant homogenates without significant cross contamination one by the other, separation of cytoplasmic side out and cytoplasmic side in plasma membrane (and tonoplast) vesicles and in the enrichment of minor cell components such as lysosomes and endoplasmic reticulum-derived transition vesicles. The procedures are facile, fast and efficient. Free-flow electrophoresis provides both analytical capabilities and the opportunity for continuous operation on a preparative scale. The procedure is nondestructive and fractions obtained retain full in vitro functional characteristics.