We have studied the cellular uptake of edelfosine (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine; ET-18-OCH3), a membrane active anticancer drug of the ether lipid family, by L1210 murine leukemia cells. Initial unidirectional linear uptake velocity was 1.1 nmol/min per 2 x 10(6) cells; at about 30 min it reached a steady-state phase of accumulation of approximately 5 nmol/2 x 10(6) cells. Concentration studies indicated no saturation kinetics from 0 to 40 microM. Studies with metabolic inhibitors displayed no energy dependence. There was no effect of chloroquine, monensin or cytochalasin B, which are known inhibitors of endocytosis. The inhibitory effect of lower temperature on uptake was moderate in extent and compatible with passive diffusion. There was no efflux of drug from preloaded cells which indicates intense binding of incorporated drug to cells. In human serum, edelfosine bound to several protein components, primarily high density lipoprotein and albumin, and this may explain why cellular uptake was slowed considerably by the presence of serum or albumin in the incubation medium. We conclude that the lipophilic ether lipid derivative edelfosine is taken up by passive diffusion by the L1210 cell. It is tightly bound to cellular structures, probably by insertion into the membrane lipid bilayer.