We examined serologically and immunohistochemically the new carbohydrate antigen CA-50 to clarify the mechanism of its high serum value and clinical significance in several liver diseases. The subjects included 145 patients with benign liver diseases and hepatocellular carcinoma (HCC). The serum CA-50 value was high in chronic active hepatitis with lobular disorganization, liver cirrhosis and HCC. It was not correlated with serum levels of GPT nor gamma-GTP. Immunohistochemical analysis revealed that proliferated bile ductules showed mainly positive staining in all subjects, whereas hepatoma cells were negative. The proliferated bile ductules with positive staining for CA-50 were quantified by an original method. The number of the proliferated bile ductules with positive staining for CA-50 was significantly correlated with the serum CA-50 value (r = 0.62, P less than 0.05). In the FPLC analysis, there was no significant difference between the expression pattern and molecular weight of CA-50 in liver diseases and pancreatic cancer. Also no difference in the carbohydrate structure that coexisted with CA-50 was detected in the ConA or LCA affinity column study. It was suggested that the increase of carbohydrate antigen CA-50 in several liver diseases might reflect the proliferation of bile ductules, and that the structure of CA-50 in benign liver diseases does not differ from that of CA-50 from patients with pancreatic cancer.