The proposed research represents a continuation of efforts to critically evaluate an immunocontraceptive approach to fertility regulation and contraceptive vaccine development based upon use of a 55K antigen (ZP3) present in porcine zona pellucida. Biological investigations include examining the longevity of the immune response and antifertility effects produced by active heteroimmmunization of squirrel monkeys with ZP3, determining minimum ZP3 dosage required to elicit zona reactive antibodies, comparing monkey immune response following ZP3 immunization with use of various adjuvants and assessing potential detrimental effects of ZP3 immunization by comparative hormonal (E2, P, LH, FSH) investigations and ovarian histological studies. Squirrel monkey immune response and antibody titers will be monitored by radioimunoassay and in vitro contraceptive potential of antisera assayed by sperm-zona binding assays. Biochemical investigations are directed toward elucidation of the chemical and structural nature of ZP3 antigenic determinants, evaluating the immunogenicity of both deglycosylated derivatives and native and synthetic peptides of ZP3, developing purification protocols for the other porcine zona glycoproteins (ZP1, ZP2, and ZP4) and initiating biochemical and immunological characterization studies on ZP1, ZP2 and ZP4 in anticipation of their future consideration for contraceptive vaccine development ZP3 and other zona components will be isolated using standard chromatographic and HPLC methodology and homogeneity verified by one-and-two dimensional acrylamide gel electrophoresis. Antigenicity and immunoreactivity of isolated components will be determined by radioimmnoassay and electrophoretic blotting. Positive findings from the proposed research would provide critical knowledge for development of a human contraceptive vaccine based upon glycoproteins present in the porcine zona pellucida.
