Primary cultures of liver cells from normal adult rats were treated with 3'-methyl-4-dimethylaminoazobenzene (3'-Me-DAB) at various concentrations for 6 days. 3'-Me-DAB treatment induced rapid proliferation of epithelial clear cells with chromosomal abnormalities and gamma-glutamyl transpeptidase (GGT) activity. In early culture, marker chromosomes were detected in 13 of 44 3'-Me-DAB-treated cultures but not in control cultures. GGT activity was not detected in the epithelial clear cells in either 3'-Me-DAB-treated or control cultures. In late culture, 21 cell lines established from 39 carcinogen-treated cultures consisted of 3 diploid cell lines, 5 pseudodiploid cell lines and 13 aneuploid cell lines. Eighteen of these 21 cell lines had marker chromosomes. Of the 2 cell lines established from 15 control cultures both were aneuploid, but a marker chromosome was detected in only one of these. GGT activity was detected in 11 of 21 cell lines established from the carcinogen-treated cultures but not in those from control cultures. Morphological features of the cell lines which varied from normal to cancerous included polymorphism, increased nuclear/cytoplasmic ratio and prominent nucleoli. No cell line established in this study developed tumors in host rats during a 1-year observation period.