A novel method for isolation and concentration of RNase T1 from Taka-Diastase is developed. It is a combination method of bentonite adsorption with dialysis desorption. In the present method, RNase T1 can be concentrated about ten-fold, the recovery of total activity was greater than 95%, and specific activity was raised 8-10 folds. Further purification with ammonium sulfate precipitation and chromatography on DEAE-cellulose and DEAE-Sephadex yields a RNase T1 which contains no pMase. pDase nor RNase T2 activities and a 750 fold increase in specific activity. Our method is more simple, rapid, and efficient than previous methods.