The effects of all-trans retinoic acid (RA) on the growth and biochemical properties of five clonal strains of neoplastically transformed rat liver epithelial cells were studied. These cell strains were derived clonally from a single line of normal diploid rat liver epithelial cells that had been transformed by treatment with N-methyl-N'-nitro-N-nitrosoguanidine. The results show that RA induces inconsistent alterations in selected phenotypic properties of these five different cell strains. Retinoic acid either depressed, enhanced or produced no effect on the colony-forming ability in soft agar, on the activity of gamma-glutamyl transpeptidase, on the amount of cell-associated fibronectin, and on the binding capacity of 125I-epidermal growth factor (EGF). The only consistent correlation observed among cell strains was between the cellular ability to grow in soft agar and the amount of cell-associated fibronectin. Enhancement of anchorage-independent growth by retinoic acid was not mediated through changes in the number of EGF receptors. Our data demonstrate that the responses to retinoic acid of clonal subpopulations of chemically transformed rat liver epithelial cells are inconsistent, even when the clonal subpopulations are derived from a common precursor.