The distribution of alkaline phosphatase was investigated in corneas of various animals by means of histochemical and biochemical methods. Special attention was paid to keratocytes that proved to be positive when a proper substrate and technique were used even if the activity differed according to animal species. Naphthol-AS-MX-phosphatate with Variamine-Blue-RT salt in the simultaneous azocoupling method was the most sensitive substrate; less suitable were naphthol-AS-phosphate, and particularly l-naphthyl-phosphate with Fast-Blue-BB-salt in the same method. Keratocytes were completely negative with β-glycerophosphate in the Gomori technique. Contrary to keratocytes, epithelial and endothelial cells were without substrate predilection. The results of both approaches showed a wide interspecies variability. The overall activity of alkaline phosphatase detected histochemically and its activity in the unsedimentable fraction of whole corneas determined biochemically was in good agreement. The highest activity was found in the calf cornea; enzyme levels decreased gradually in the bovine, rabbit, dog, sheep, and pig cornea