Gangliosides are immunosuppressive to many classes of immune cells, and shedding of these glycosphingolipids by tumour cells may regulate immune responses in cancer, and protect tumours from host immune destruction. One mechanism of immunosuppression by gangliosides in vitro involves competition with interleukin-2 receptors (IL-2R) for binding of IL-2. Previous studies on inhibition of IL-2-mediated events by gangliosides have been conducted in the presence of high levels of fetal bovine serum (FBS). However, gangliosides shed by tumours in vivo will encounter immune cells in the low protein microenvironment of the tissue fluid. In order to better mimic physiological conditions, we have examined immunosuppression by gangliosides towards IL-2-dependent HT-2 cells in a low serum-low protein medium. The ability of gangliosides to inhibit IL-2-stimulated DNA synthesis in HT-2 increased dramatically as the serum concentration in the culture medium was decreased; the 50% inhibitory concentration (IC50) value for GM1 was 13 microM under low serum conditions, 14-fold lower than the value obtained in 10% FBS. Further investigation revealed that the mechanism of immunosuppression by gangliosides in low serum-low protein medium involved interference with the IL-2/IL-2R system. Ganglioside-mediated inhibition was dependent on the continued presence of the glycolipids during the first few hours after IL-2 stimulation, and could be reversed by increasing levels of IL-2. Receptor binding experiments demonstrated that gangliosides blocked the interaction of IL-2 with high-affinity IL-2 receptors on HT-2. Taken together, these results support the view that gangliosides will act as much more potent suppressors of IL-2-dependent processes in vivo in the vicinity of a tumour.