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In the United States, a large percentage of patients with hepatocellular carcinoma are serologically negative for hepatitis B. We conducted a retrospective study to determine the prevalence of hepatitis C antibody in the sera of 59 patients with hepatocellular carcinoma who were HBsAg-negative and had no evidence of alcoholic liver disease, primary biliary cirrhosis, autoimmune hepatitis, hemochromatosis or alpha 1-antitrypsin deficiency. Twenty patients (34%) were hepatitis C antibody-positive and hepatitis B core antibody-negative. All twenty patients had underlying cirrhosis, and seven (35%) had histories of transfusions. Eleven (19%) additional patients were also hepatitis C antibody-positive but were hepatitis B core antibody-positive as well. Twenty-one (36%) patients were both hepatitis C antibody- and hepatitis B core antibody-negative and seven (12%) were hepatitis C antibody-negative but hepatitis B core antibody-positive. The prevalence of hepatitis C antibody was also determined among three other population groups serving as controls and found to be 14% in 28 HbsAg-positive patients with hepatocellular carcinoma, 44% in 76 patients with cryptogenic cirrhosis and 0.5% in 200 consecutive volunteer blood donors. We conclude that hepatitis C antibody is prevalent among patients with hepatocellular carcinoma and may therefore be a common causative agent of this disease. A significant number of patients with and without cirrhosis, negative for hepatitis C antibody and hepatitis B core antibody, remain without a discernible cause for this malignancy. Perhaps a second- or third-generation test will detect hepatitis C antibody in some of these patients.