Preparative polyacrylamide gel electrophoresis has been used to purify the enterotoxin of Clostridium perfringens from Sephadex G-100 extracts. Purified toxin of high specific activity was eluted in 1 to 3 h, depending upon the length of the acrylamide gel used. Recovery of biological activity with this technique ranged from 80 to 90%. The purity and physical characteristics of the toxin were similar to those previously reported for the protein purified by other methods. Use of preparative electrophoresis will enable the production of larger amounts of high-specific-activity toxin in a shorter time than other currently available procedures. This method was also used to isolate a form of enterotoxin that has a mobility, relative to bromophenol blue tracking dye, of 0.87 to 0.90 in 7% acrylamide gels.