Antibodies against constitutive proteins of different types of intermediate-sized filaments were used in immunofluorescence microscopy on frozen sections of normal rat liver and various rat liver tumors induced by treatment with nitrosamines. Antibodies to tonofilament prekeratin stained bile duct epithelia and hepatocytes of normal liver and hepatocellular carcinoma cells and ductal cells of cholangiofibromas. These cells were not significantly stained by antibodies to vimentin. By contrast, antibodies to vimentin stained mesenchymal cells of normal liver and cells of early and advanced angiosarcomas and of undifferentiated spindle cell sarcoma. These mesenchymal tumor cells were not stained with antibodies to prekeratin. (5,6,8,(11)(12)(13) has stimulated our interest in the possible maintenance of such molecular markers in tumors grown in the body.As a first example we have used the liver (in which different types of tumors, such as adenomas, hepatocellular carcinomas, cholangiofibromas, cholangiocarcinomas, angiosarcomas, and fibrosarcomas can occur) to determine whether epithelial and mesenchymal tumors could be distinguished by the use of antibodies to different filament proteins. The search for a molecular marker for identification of mesenchymal tumors of liver was especially desirable because mesenchymally derived liver tumors are important in human (14) and experimental animal (15) pathology. Reports of a high incidence of angiosarcomas in workers exposed to inorganic arsenic and gaseous vinyl chloride and in patients treated with Thorotrast or certain steroid hormones have increased the interest in this tumor (14, 16). The structure and morphogenesis of vascular liver tumors in humans and experimental animals appear to be largely identical (15,17). In both humans and animals, differential diagnosis of epithelial and mesenchymal liver tumors is sometimes difficult. A clear distinction, however, would be important not only for prognostic appraisals in human pathology but also for evaluation of animal experiments such as dose-response studies of carcinogenesis induced by chemical compounds (18, 19).In this study we show that immunofluorescence microscopy with antibodies to two molecular markers, prekeratin and vimentin, allows the character and origin of epithelial and mesenchymal cells to be distinguished in neoplastic liver cells and thus facilitates the differential diagnosis of liver tumors.MATERIALS AND METHODS Animals. Liver tumors were induced in male SpragueDawley rats (n-200 g body weight at the beginning of treatment) by oral treatment with nitrosamines (for methods, see ref. 20). Seven hepatocellular carcinomas, 3 cholangiofibromas, 10 angiosarcomas, and 1 undifferentiated spindle cell sarcoma were examined. Four of the hepatocellular carcinomas had developed in animals treated with 50 mg of N-nitrosomorpholine (in a total of 100 ml of their drinking water) for 3 weeks and killed between weeks 52 and 77 after withdrawal of the carcinogen. (Three of these carcinomas occurred in asso...