AbstractLeukemia inhibitory factor (LIF) is a cytokine previously shown to maintain pluripotent embryonic stem cells in their undifferentiated state. We have examined the effects of LIF in nullipotent embryonal carcinoma cell lines, and have found that LIF blocks differentiation induced by retinoic acid and at low temperature in OTF9 cells. LIF did not block differentiation in a parent F9 cell line. For OTF9 cells, LIF acts early in differentiation, inhibiting the appearance of parietal endoderm-type product cells. However, it acts subsequent to retinoic acid, and at least one early retinoic acid-induced event is unaltered in the presence of LIF. This finding provides both a means of dissecting the cascade of events leading to EC cell differentiation, and a well-characterised target cell type for studying the mechanism of action of LIF.