A concanavalin-A(Con A)-resistant variant of the pluripotent mouse embryonal carcinoma cell line, PSA1-NG2, was isolated. This variant, designated NG2-2.16, fails to exhibit the extensive spontaneous differentiation displayed by PSA1-NG2 in colonies in vitro and in tumours in vivo. The molecular nature of the defect in NG2-2.16 cells was not revealed by quantitative studies of the binding, uptake and metabolism of tritiated Con A, or by Western blotting of membrane and whole cell homogenates, thus indicating the defect to be the result of a more subtle molecular alteration. Statistical evidence suggests that the same mutation is responsible for both the Con A resistance and the lack of spontaneous differentiation. NG2-2.16 cells were induced to differentiate by exposure to retinoic acid, suggesting that the mutation affects the regulation of differentiation rather than the potential for differentiation.