Molecular nature of a sperm acrosomal antigen recognized by HS-13 monoclonal antibody

Abstract

Among the numerous monoclonal antibodies generated against human sperm antigens, HS-13 monoclonal antibody was shown to react with an intra-acrosomal antigen from human, mouse and rat. In this study, HS-13 was used as the affinity ligand for the purification of the cognate antigen from human sperm by immunoaffinity chromatography. The purified cognate antigen from human sperm, designated as HSAg-13, was found to be a protein with a molecular weight of approximately 80 kDa on SDS-PAGE in the presence of reducing reagents. This monoclonal antibody was used as the probe to study the tissue distributions and developmental expression of the cognate antigen from human, mouse and rat by immunohistochemical assays. It was concluded that the antigen recognized by HS-13 antibody is highly sperm specific and found only in sperm and mature testis, but not in any other somatic tissues examined in human and mouse. The antigen was shown to be expressed at the postmeiotic stages of spermatogenesis in mouse and rat. By using indirect immunofluorescent staining assay, HS-13 was shown to react only with the methanol-fixed acrosome-intact sperm but not with the live sperm. Following calcium ionophore A23187 treatment, acrosome-reacted sperm showed either negative staining or residual staining in the equatorial region, suggesting the intra-acrosomal location of HSAg-13. The spontaneous acrosome reaction following overnight incubation in BWW medium resulted in a statistically significant decrease of antibody-stained human sperm. In view of excellent correlations for the scoring of acrosome-intact sperm with that of fluorescence-labeled Pisum sativum agglutinin (PSA) probe, HS-l3 monoclonal antibody can be routinely used for monitoring sperm capacitation and acrosome reaction.