A strategy for identifying candidate sperm antigens for immunocontraception: isolation of human testis cDNA clones using polyclonal antisera directed against hamster acrosomal membrane preparation.
Components of the mammalian sperm acrosome that have been conserved during evolution are probably essential for fertilization and are therefore potential antigens for the development of an immunocontraceptive vaccine. In order to identify such protein components, a series of specific polyclonal antisera were generated by immunizing rabbits with purified acrosomal membrane fractions from hamster epididymal spermatozoa. Antisera were finally selected using immunological and in-vitro fertilization assays, and used to then screen a human testis lambda gt11 cDNA library. As a result of this screening over 70 clones were identified, selected and purified. The cDNAs were amplified by polymerase chain reaction (PCR) and the inserts characterized by restriction enzyme digestion and oligonucleotide probing techniques. The functional activity beta-galactosidase fusion proteins expressed by these clones (HA5-2, HA6-2 and HB4-1) inhibited significantly fertilization and reduced spermatozoa binding compared to controls. To date, sequence data has been obtained from HB4-1 (1.75 kb). The first 1132 nucleotides displayed > 96% homology to human testis-specific lactate dehydrogenase (LDH-C4) gene, the product of which is a known candidate antigen for a contraceptive vaccine. This finding suggests that a strategy involving the screening across species for conserved moieties of the mammalian acrosome may be useful for identifying candidate antigens for immunocontraception.
Adoyo
P A
PA
Institute of Primate Research, Nairobi, Kenya.
Moore
A
A
Moore
H D
HD
D000177:Acrosome / Q000276:immunology*; D000818:Animals; D000906:Antibodies; D000941:Antigens; D000954:Antigens, Surface / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D003001:Cloning, Molecular; D003267:Contraception / Q000379:methods*; D006224:Cricetinae; D004262:DNA Restriction Enzymes; D018076:DNA, Complementary; D004822:Epididymis; D005260:Female; D005307:Fertilization in Vitro; D006801:Humans; D007527:Isoenzymes; D007770:L-Lactate Dehydrogenase / Q000235:genetics / Q000276:immunology; D008297:Male; D008647:Mesocricetus; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D011817:Rabbits / Q000276:immunology; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013094:Spermatozoa / Q000166:cytology / Q000276:immunology*; D013379:Substrate Specificity; D013737:Testis / Q000276:immunology*
D000906:Antibodies; D000941:Antigens; D000954:Antigens, Surface; D018076:DNA, Complementary; D007527:Isoenzymes; D007770:L-Lactate Dehydrogenase; D004262:DNA Restriction Enzymes