The expression of human leukocyte antigen (HLA) on human ejaculated sperm has been investigated by using cellular binding radioimmunoassay (CB-RIA) with sperm samples purified by Percoll discontinuous gradient and monoclonal antibodies specific to the frame work of HLA molecules. The results in packed sperm obtained from ten healthy males indicated that HLA class II and its subclasses, DP, DQ, and DR, were expressed at a low level on human ejaculated sperm, while HLA class I was not detected. Furthermore, we could not detect the individual variation of HLA expression on ejaculated sperm in the cases of twenty healthy males. K562, which is B lymphoid cell line and lacks HLA class I molecules, showed detectable amounts of HLA class I molecules after exposure to interferon gamma (IFN-gamma) and/or tumor necrosis factor alpha (TNF-alpha). However, the treatment of sperm with IFN-gamma and/or TNF-alpha had no effect on the expression of HLA class I and class II molecules.