Anti-SLIP1-reactive proteins exist on human spermatozoa and are involved in zona pellucida binding

Sulpholipid immobilizing protein 1 (SLIP1) is an evolutionarily conserved 6 8 kDa plasma membrane protein, present selectively in germ cells. We have p reviously shown that mouse sperm SLIP1 is involved in sperm-zona pellucida (ZP) binding. In this report, we extended our study to the human system. Im munoblotting demonstrated that anti-SLIP1-reactive proteins (mol, wt 68 and 48 kDa) could be extracted from human spermatozoa by an ATP-containing sol ution, a result that is consistent with observations in other species. Dire ct immunofluorescence, using Cy3-conjugated anti-SLIP1 IgG, revealed SLIP1 staining over the acrosomal region, with higher intensity at the posterior area. Using the human sperm-ZP binding assay, we demonstrated that pretreat ment of human spermatozoa from three donors with anti-SLIP1 IgG revealed lo wer numbers of zona-bound spermatozoa, as compared to the corresponding con trol spermatozoa treated with normal rabbit serum IgG, This decrease in zon a pellucida binding was not from an antibody-induced decline in sperm motil ity or an increase in the premature acrosome reaction. The results strongly suggest that anti-SLIP-reactive proteins on human spermatozoa play an impo rtant role in ZP binding.