Electroblotting of Proteins from 2-D Polyacrylamide Gels

The ability of two-dimensional electrophoresis (2-DE) to separate complex mixtures of proteins, such as represented by cells, tissues, and even whole organisms, has been recognized for more than 20 yr (1 ). Using “standard”-format (around 20�20 cm) 2-D gels, the method is capable of routinely separating 2000 proteins from whole-cell and tissue extracts. The resolution capacity can be extended significantly (up to 5000–10,000 proteins) using large-format (40�30 cm) 2-D gels (2 ). Over the last 20 yr, 2-DE has been used primarily as an analytical tool for the characterization of proteins by their charge (pI ), size (M r ), and relative abundance. Specialized computer software has been developed for the qualitative and quantitative analysis of 2-DE protein patterns (3 ), and these systems have been used to construct several comprehensive databases of protein expression in a variety of cell types and tissues (for examples, see ref. 4 ).