Monoclonal antibody AG7 inhibits fertilization post sperm-zona binding

Monoclonal antibodies (mAbs) against sperm cells are currently being used in an effort to define spermatozoal antigens involved in the fertilization process. We have produced a number of anti-human sperm mAbs by immunization of female mice with the 100,000 x g supernatant of octylglycoside-solubilized washed human sperm. From a panel of mAbs, 1 antibody, AG7, was selected and characterized due to its fertilization-inhibiting characteristics. MAb AG7 defines a sperm acrosome antigen-1 (SAA-1) located in the acrosomal region of human sperm as evaluated by indirect immunofluorescence. Staining of life sperm cells indicated that the antigen is present on the sperm surface. SAA-1 was also found on sperm of several other mammalian species, implying evolutionary conservation of the antigen. SAA-1 was first observed on testicular sperm and can be followed through epididymal transit, ejaculation, and capacitation. When applied in a mouse in vitro fertilization assay, mAb AG7 inhibits fertilization by greater than 95%, and inhibition is dose dependent, with half-maximal inhibition at 0.8 micrograms/ml. The block to fertilization could not be attributed to sperm agglutination, inhibition of motility, interference with adhesion to the zona pellucida, or inhibition of fusion with the oocyte membrane. MAb AG7 was demonstrated to inhibit calcium influx in spermatozoa in vitro (measured using the fluorescent indicator fura 2), a prerequisite for the acrosome reaction. Initial biochemical characterization of the antigen suggests it is proteinlike in nature, with a molecular weight of approximately 220 kD. The results suggest that SAA-1, identified by mAb AG7, is a sperm antigen crucially involved in the fertilization process, possibly an atypical steroid receptor or ion channel located within the sperm plasma membrane.