The established mouse cell line NIH 3T3 has been used with considerable success over the past three years as the basis of an in vitro transformation assay for demonstrating the presence of transfectable transforming genes in the DNA of certain human and rodent tumour cells (for review see ref. 1). In the case of the human bladder carcinoma cell lines EJ and T24, this approach has led to the molecular cloning of a transforming gene which is closely related to the rat-derived Harvey sarcoma virus oncogene, v-Ha-ras. A single point mutation, which distinguishes these genes from their normal human homologue (c-Ha-ras1), is thought to be solely responsible for their transforming potential. However, carcinogenesis in both humans and laboratory rodents is a multi-stage process (reviewed in ref. 11) of which the NIH 3T3 cell, already partly transformed, may represent only the penultimate stage. We therefore chose to examine the transforming effects of the EJ oncogene in a hamster fibroblast system originally developed in our laboratory to study stages in carcinogen-induced malignant transformation of normal diploid cells. We show here that EJ c-Ha-ras-1 lacks complete transforming activity when transfected into normal fibroblasts which have a limited lifespan, but can fully transform fibroblasts that have been newly 'immortalized' by carcinogens.