The human sperm protein SP-10 was previously defined as a "primary vaccine candidate" by a World Health Organization Taskforce on Contraceptive Vaccines. By one- and two-dimensional immunoblots, we show that SP-10, extracted from ejaculated human sperm, demonstrated a polymorphism of immunogenic peptides from 18 to 34 kDa, a pattern that was conserved from individual to individual and was not altered by reducing agents. The majority of the antigenic peptides possessed isoelectric points of approximately 4.9. Immunocytochemistry on testis sections indicated that SP-10 was localized to round spermatids and spermatozoa within the adluminal compartment of the seminiferous epithelium. Immunofluorescence showed that SP-10 was not associated with the surface of acrosome-intact, ejaculated sperm. Light and electron microscopic immunocytochemistry localized SP-10 throughout the acrosome, and electron microscopic evidence demonstrated a bilaminar array in association with the inner aspect of the outer acrosomal membrane and the outer aspect of the inner acrosomal membrane. After induction of the acrosome reaction with the ionophore A23187, SP-10 remained displayed on the sperm head in association with the inner acrosomal membrane and equatorial segment. The results indicate that the MHS-10 monoclonal antibody may be used as a marker of acrosome development in the human and as a probe to evaluate acrosome status. The results also support the hypothesis that inhibition of sperm-egg interaction by anti-SP-10 monoclonal antibody may occur as a result of antigen exposure following the acrosome reaction.

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This study characterizes a common antigen recognized by two monoclonal antibodies (mAbs) that immunoreact with the principal piece of the human sperm flagellum. By means of immunofluorescence microscopy, mAbs S69 (IgM) and S70 (IgG1) (WHO Task-force nomenclature) were observed to bind to the principal piece of methanol- or detergent-permeabilized human spermatozoa, but did not react with live swimming spermatozoa as assessed by immunofluorescence microscopy. Faint immunofluorescence was also seen on the connecting piece in approximately 40% of the spermatozoa. Immunoreactivity in both regions was resistant to sequential extraction with Triton X-100, sodium thiocyanate, and urea. Pre-embedding electron microscopic immunogold labeling of ejaculated spermatozoa with mAb S69 showed gold particles located on the fibrous sheath. Immunoreactive peptides of 68, 53, and 45 kDa were recognized by both S69 and S70 mAbs on immunoblots of nonreduced human sperm extracts, while a 68-kDa band and a strongly immunoreactive triplet from 54 to 51 kDa were recognized in reduced sperm extracts. Human fibrous sheaths were isolated by differential solubilization and centrifugation and characterized by transmission electron microscopy. The 68-kDa and 54-51-kDa bands were enriched and found to be major polypeptides in the isolated fibrous sheath fraction. These results suggest that the S69/S70 antigen, which we term SP (sperm protein) (68 kDa, 54-51 kDa), is a component of the human fibrous sheath.