Six human pancreatic cancer cell lines PK-1, -8, -9, -12, -14 and -16, were established. They originated from either primary pancreatic cancer biopsy or liver metastasis biopsy, or xenografts of these biopsy specimens in athymic nude mice. The primary tumors were all well differentiated adenocarcinomas of pancreatic duct origin. The six established PK cell lines were all CEA positive and had tumorigenicity in athymic nude mice. Morphology of the xenografted tumors was closely similar to that of the original tumor. PK cells grew slowly with the doubling time of 41.3 to 82 hr and showed aneuploid chromosome pattern. High levels of glucose-6-phosphate dehydrogenase (G6PDH) and lactic dehydrogenase (LDH) were found in each cell extract. Trypsin was not detected in cell extracts except PK-8 and PK-9. In chemosensitivity test, all of PK cell lines were sensitive to aclacinomycin A (ACM), and PK-1 and PK-8 were sensitive to 5-Fluorouracil (5-Fu) at concentrations of 0.02 microgram/ml, ACM and 1 microgram/ml, 5-Fu, when the drugs were used for over 48 hr. At higher concentrations, they showed time independent sensitivity to mitomycin C (MMC). PK-9 was resistant to 5-Fu and MMC.