IMMUNOLOGICAL IDENTIFICATION OF ZONA-PELLUCIDA-2 (ZP2) PROTEIN IN HUMAN OOCYTES

The ZP2 protein is a zona pellucida glycoprotein that plays a major ro le in fertilization. It mediates secondary binding of spermatozoa and is one of the proteins that are involved in zora 'hardening'. ZP2 prot eins were identified in various mammalian zonae pellucidae. Their prim ary structures are highly conserved as revealed by cDNA cloning. Antis era were used against synthetic peptides generated either against a ZP 2 amino acid that is homologous in human and mouse ZP2 amino acid sequ ences (AS ZP2-20) or antibodies against a synthetic human ZP2 peptide (AS ZP2-26). Immunoblots showed that antiserum AS ZP2-20 and AS ZP2-26 strongly recognized human ZP2 protein with an apparent molecular mass of about 72 kDa; both antisera reacted with a minor immunoreactive po lypeptide at 96 kDa. In human ovary sections, both antisera revealed i mmunoreactivity to human zonae pellucidae. Immune-electron microscopy demonstrated an equal distribution of ZP2 throughout the human zona pe llucida. Considerable amounts of immunoreactive material were observed in the ooplasm; some ramification-like extensions of zona pellucida a ntigen were found close to cells surrounding the oocyte. Our results i ndicate that antisera against synthetic ZP2 peptides can be used as sp ecific markers for the identification of ZP2, protein in human oocytes .