Abstract
A modification of O'Farrell's method of two-dimensional polyacrylamide gel electrophoresis has allowed for the resolution of erythrocyte membranes showing up to 200 individual components. Data is presented which indicates that this protein heterogeneity is not produced by artifactual protein-protein aggregation, endogenous protease activity or secondary charge modification. Similar patterns are obtained when the samples are added to the unpolymerized isoelectric focusing gel, and isolated and stored in protease inhibitor. Individual spots could be eluted off of stained gels, resolubilized under extreme detergent solubilization conditions and run on one-dimensional gels; these run as individual bands. One of the advantages of the method is the presence of sodium dodecyl sulfate in the solubilization procedure. The method chosen for solubilization prior to isoelectric focusing appears to cause selective aggregation of all or most of the spectrin and band 3 proteins. This further allows for excellent resolution of minor components.
