Recently, cholangiocellular carcinoma (CCC) was successfully induced in the hamster by infecting with Clonorchis sinensis following hepatocarcinogen treatment and has been proposed as a suitable model to study the pathogenesis of human CCC. In this hamster model, oval cells are suggested to be cells of origin of CCC. More direct analysis of histogenesis of CCC would become possible if large numbers of highly purified oval cells of hamster origin are obtained. In this study, we describe successful isolation of highly purified oval cells from hamsters. Oval cells were induced by diethylnitrosamine and 2-acetylaminofluorene treatment under choline deficient diet and isolated by centrifugal elutriation method. This isolated cells were highly homogenous in size (10.9+/-1.1 microm in diameter) and had a high nuclear to cytoplasmic ratio, an oval-shaped nucleus and a few cytoplasmic organelles. Immunocytochemically, 85.4+/-1.6%, 75.1+/-2.0%, 62+/-1.5% and 25.6+/-2.7% of the isolated cells were positive for cytokeratin 19, OV-6, albumin and alpha-fetoprotein, respectively, indicating that these cells had phenotypic characteristics of both hepatocytes and bile duct epithelium. The isolated cells were therefore considered to be hamster oval cells.