Activities of acid and alkaline phosphatases were examined in spermatozoa isolated from 177 semen samples differing in sperm counts. Alkaline phosphatase was also determined in seminal fluid. The enzymes were assayed using disodium p-nitrophenyl phosphate as substrate and were studied with respect to susceptibility to various concentrations of tartrate (acid) and to heat (alkaline). Electrophoretic separation of alkaline phosphatase from seminal fluid was performed using an Helena apparatus. The results showed that acid phosphatase activity in spermatozoa decreased with increase in sperm densities and that elevation of tartrate from 0.028 to 0.17 M usually correlated an inhibition of the enzyme from 72% to 78% (mean values). Alkaline phosphatase was very low in sperm and generally below the sensitivity of the method used. Activity of alkaline phosphatase in seminal fluid showed a tendency to increase with the increase in sperm counts, but the significance of differences between groups was not statistically valid. Exposure of seminal fluid to 55 degrees C for 16 min resulted in enzyme inactivation of about 90% and in this respect the alkaline phosphatase resembles the enzyme of bone origin. The electrophoretic pattern, however, did not confirm this view and the type of alkaline phosphatase in seminal fluid is not clear.