A human gastric carcinoma cell line, KATO-II, and its subline, KATO-III, have been established in vitro from a pleural effusion of a 55-year-old male patient. Kato-II cells have been maintained using a culture medium containing human cord serum and KATO-III cells have been cultured with a medium containing fetal bovine serum. Both cell lines are morphologically quite similar; they grow in vitro floating free and have cytological features of signet ring cells. Population doubling times for KATO-II and KATO-III were 74 hours and 36 hours, repsectively. The modal chromosomal numbers of these cell lines fell in a tetraploid range. Heterotransplantation of KATO-II and KATO-III cells was successfully done in the cheek pouches of antithymocyte serum-treated hamsters. Microscopical appearance of the resultant tumors was consistent with poorly-differentiated adenocarcinoma comparable to the original tumor.