Ectopic production of salivary-type amylase was demonstrated in a human gastric carcinoma cell line (KMK-2) maintained in vitro for more than 3 years. Electron-dense granules, which appeared as zymogens, were observed in the tumor tissue, in cancer cells in the peritoneal fluid (from which the present cell line had been derived), and in cultured cells in early passages. These granules, however, decreased and gradually disappeared during cultivation. Although such a morphologic alteration was recognized, the property of amylase synthesis has been maintained. The presence of alpha-amylase in the cultured cells was demonstrated cytochemically by the immunoperoxidase method. The enzyme secreted and accumulated in the culture medium was partially purified and characterized. Alpha-amylase of KMK-2 cells closely resembled salivary-type amylase in gel filtration profile and disk gel electrophoresis. Immunologic cross-reaction was observed between these enzymes. Secretion into the medium was constant, and the enzyme concentration in the cytoplasm was relatively high when the cells had reached confluence. Prednisolone increased the amylase production two-fold in the cells.