Preparative free-flow electrophoresis, a versatile technology complementing gradient centrifugation in the isolation...

Free-flow electrophoresis (FFE) exploits differences in the overall charge of bio-particles separating cells, organelles, macromolecules, ions, etc. according to their distinct electrophoretic mobility and isoelectric point (pI) values. Indeed, around a neutral pH organelles usually exhibit a negative surface charge, migrating in an electric field from the cathode toward the anode. Since its introduction more than five decades ago by Barrollier et al., Z. Naturforsch. 1958, 13b, 745-755 and Hannig, Z. Anal. Chem. 1961, 181, 244-254, FFE has become an established analytical and preparative separation method for the isolation of a variety of organelles. Particularly, in sophisticated, multistep separating processes to separate subpopulations of organelles, it has gained, meanwhile, a position as a versatile technology and essential element. Relying on the distinct surface charges instead of buoyant densities of cell organelles, the FFE technology is best supporting a preceding centrifugation-based fractionation of subcellular compartments in the second dimension. In the following review, the two-step isolation and purification of subpopulations of classic animal and plant cell organelles will be mainly exemplified.