A series of closely related mouse fibroblast cell lines that differ in their content of neutral ether-linked glycerolipid and fatty acids has been used to investigate the relationship between lipid composition and tumorigenicity. Although these cell lines, derived from the same parental culture, were selected without reference to transformation or tumorigenicity, their ability to form tumors in irradiated mice was found to be closely correlated with ether-lipid content. The cell line with the highest level of ether-lipid (designated F40) produces more tumors, the tumors appear more rapidly than when parental cells are injected, and the number of F40 cells required for tumor induction is less by a factor of approximately equal to 1000. F40 tumors are highly invasive, readily metastasize, and rarely regress, in contrast to the occasional benign tumors produced by the parental cell line. Cell lines that are intermediate in their lipid composition appear to be intermediate in tumorigenicity. This panel of graded cell lines provides a useful model system for both in vitro and in vivo studies on the acquisition of tumorigenicity and malignancy in cultured cells.