Esterases in nine rat hepatic and hepatoma-derived cell lines and normal rat liver homogenate were detected by polyacrylamide gel electrophoresis coupled with active staining with alpha-naphthyl acetate or butyrate as a substrate. The esterase band patterns of the non-cancerous and oncogene-transformed cell lines were alike, but different from those of hepatoma cell lines and normal rat liver homogenate. The former groups of cells might have completely lost the characteristics of rat liver parenchymal cells, or else they might have their origin at cells other than liver parenchyma. The esterase patterns of the hepatoma cell lines (e.g., McA-RH7777) and the normal rat liver highly resembled with each other, exemplifying the slight biochemical deviation of cancer from normal cells. However, two-dimensional electrophoretogram for the McA-RH7777 cell line showed a prominent esterase spot (pI 6.0-M(r) 110 kDa) that was lacking in the normal liver. This result indicates that there is invariably some change in esterase expression between the cancer cells and the normal liver cells.