Two-Dimensional Polyacrylamide Gel Electrophoresis of Proteins

Abstract

Since O’Farrell (1) introduced the improved technique for high resolution two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), it has become one of the most powerful tools for the separation and quantification of proteins from complex mixtures. The principal reason for this is that the method employs separation of denatured proteins according to two different parameters, molecular weight and isoelectric point. Consequently, it has sufficient resolution to separate individual proteins as discrete spots on the gel. Each parameter may also be varied and therefore, with the modification of non-equilibrium pH-gradient electrophoresis (NEPHGE) to analyze basic proteins (2), almost any polypeptide may be investigated. Thus to date, the O’Farrell 2-D gel system has no serious rivals, with the possible exception of the Kaltschmidt and Wittmann (3) gel system for analyzing ribosomal proteins. Ribosomal proteins, however, may be adequately separated with NEPHGE.