A variety of conditions of cryopreservation were evaluated in order to define a single procedure for freezing the amebae of pathogenic Naegleria and Acanthamoeba. The average best conditions for freezing the three species studied were; lxlO1’ exponentially growing amebae/ml of freezing medium consisting of 12% dimethylsulfoxide, 20% heat-inactivated bovine calf serum, 4% glucose, in Mix ameba medium; 30 min equilibration at 23” С (room temperature), followed by 60 min at -20° C, with storage at -70° C. Under these conditions viability after 1 month of freezing was 53% for Acanthamoeba castellami, 64% for Naegleria fowleri, and 66% fot Naegleria australiensis. After 12 months of freezing, viability was 39% for A. castellami, 47% for N. fowleri, and 53% for N. auslraliensis.