ANTIBODY-DIRECTED AGAINST PLASMA-MEMBRANE COMPONENTS OF EQUINE SPERMATOZOA INHIBITS ADHESION OF SPERMATOZOA TO OVIDUCT EPITHELIAL-CELLS IN-VITRO

Before fertilization, equine spermatozoa adhere to oviduct epithelial cells (OEC) of the mare. The biochemical basis for this adhesion has n ot been determined. Our objective was to produce an antiserum to block this interaction. Ejaculated spermatozoa were subjected to nitrogen c avitation and spermatozoal plasma membranes enriched by sucrose densit y gradient centrifugation; membrane enrichment was confirmed by compar ative alkaline phosphatase analysis, electron microscopy, and one- and two-dimensional PAGE. Periacrosomal plasma membrane was used as an im munogen for the production of an antiserum, which recognized several c omponents of spermatozoal plasma membrane on Western blots. Antigen-bi nding fragments (Fab) were isolated by papain digestion from a specifi c antiserum and from nonimmunized rabbit IgG (control). The periacroso mal regions of epididymal and ejaculated spermatozoa were immunolabele d with antiserum Fab but not control Fab. The immunoneutralizing activ ity of antiserum Fab was tested in fluorescent cell-binding assays by competitive inhibition of the binding of spermatozoa to OEC monolayers or explants. In both assays, binding of spermatozoa to OEC was reduce d as the concentration of specific Fab increased. These results sugges t that one or more protein or glycoprotein components of the rostral s permatozoal plasma membrane mediate adhesion between spermatozoa and o viduct epithelium in vitro.