Immobilized pH 2.5–11 gradients for two‐dimensional electrophoresis

Abstract

Extremely wide immobilized pH gradients, pH 2.5–11, for isoelectric separation of complex protein mixtures are described. These pH gradients are theoretically and practically the maximum that can be achieved at present with the available acrylamido buffers and titrants. Conditions are described for reducing conductivity and electroendosmosis in extreme pH ranges. Furthermore, new conditions are described for the separation of proteins in the second dimension. Using this protocol, nearly all the possible cellular products can be separated in one single two‐dimensional map.