The broad objective of this application is to identify molecules on the surface of guinea pig spermatozoa that are involved in recognizing and adhering to the zona pellucida of the egg. The rationale behind the proposed experiments stems from the established observation that there are two potent inhibitors of sperm-zona binding: 1) anti-sperm antibodies and 2) the fucoidin polysaccharide. The goal will be to identify pertinent receptors or targets from sperm that recognize these two probes. To exploit the antibody approach, a rabbit anti-guinea pig sperm antibody will be raised whose F'ab fragments inhibit sperm-zona adhesion. After solubilization of material from sperm or testis, the antigen(s) involved in this inhibition will be progressively isolated and identified by assaying for their ability to """"""""neutralize"""""""" the F'ab inhibition. Techniques proposed for this task include immunoaffinity chromatography, acrylamide gel electrophoresis, and classical permeation, ion-exchange, or hydrophobic chromatography. As a complementary approach, fucoidin-binding material from sperm or testis extracts will be isolated using fucoidin-sepharose affinity chromatography. This material will be used to determine whether the functional antibody targets are the same or different from the fucoidin receptors on sperm. Information gained from this work will help to elucidate the molecular basis of mammalian fertilization. Identification of functional sperm surface antigens will provide clues to the basis of immunological infertility and may suggest a specific approach to a fertility vaccine. Identification of functional surface components which bind to fucoidin will shed light on conserved mechanisms of gamete recognition in particular and cell-cell adhesion in general.
