Determination of protein molecular weights by gel permeation high pressure liquid chromatography.

An essential part of the characterization of any protein is the determination of its molecular weight. The method of choice for these determinations, because of its simplicity and rapidity, has most frequently been sodium dodecyl sulfate (SDS) gel electrophoresis (See Chapter 6 ). The alternative method of gel filtration under denaturing conditions (1,2) has not been so widely used, in part as a result of the longer times required for a single run. However, recent developments in gel filtration supports for high pressure liquid chromatography (HPLC) have made more rapid separations possible, and thus gel permeation HPLC is becoming a widely used technique for molecular weight determinations (3,4). Gel permeation HPLC, in addition to taking less time than SDS gel electrophoresis, allows easier quantitation and recovery of separated proteins, and the resolution is better than that achieved by gel filtration with conventional materials.