A simple and rapid method, using only two chromatographic steps, is described for the purification and preparation of gamma gamma enolase isoenzymes from human and beef brain extracts. In the first step, a crude gamma gamma enolase was obtained by chromatography on Q-Sepharose Fast Flow column. The crude fraction was then purified by high performance anion exchange chromatography on a Mono-Q column. gamma gamma enolase obtained in this manner was shown to be homogeneous by two dimensional polyacrylamide gel electrophoresis and by high performance gel permeation chromatography. The yield of gamma gamma enolase by this method was 7-8 mg of pure enzyme per 100 g of brain.