Studies with Synthetic Peptides of 80 kDa Human Sperm Antigen (80 kDa HSA).
OBJECTIVE
The 80 kDa human sperm antigen (HSA) is a sperm-specific and conserved antigen, capable of inducing immunological infertility. Partial N-terminal amino acid sequences of 80 kDa HSA (Peptide NT) and its peptides obtained by digestion with endoproteinase Lys-C (peptides 1-4) and endoproteinase Glu-C (peptides 5-6) did not show any sequence homology with reported known proteins deposited in the Gen-Bank. These sequenced peptides were synthesized and conjugated to key hole limpet haemocyanin (KLH) and evaluated for its antifertility effects. The present communication describes the characterization of these peptides and their antibodies.


METHODS
Peptides NT, 1, 2, 3 and 4 were synthesized and conjugated to KLH. Antibodies to KLH conjugated peptides were raised in rabbits by active immunization and the antibody titer was determined by enzyme-linked immunosorbent assay (ELISA) using sperm extract coated wells. The binding specificity of the synthetic peptides or purified 80 kDa HSA to their antibodies was assessed in the presence of various doses of respective synthetic peptides or 80 kDa HSA. The binding specificity was further confirmed by Western blot analysis. Antipeptide antibodies were also checked for sperm agglutinating activity, in-vitro.


RESULTS
Active immunization of rabbits elicited significant antibody titers against the synthetic peptides, except for peptide 3. Antipeptide antibodies specifically recognized the native protein in an ELISA and induced in-vitro agglutination of human, rat and monkey sperm. In addition, Western blot analysis showed that these antipeptide antibodies specifically bind to the 80 kDa HSA band of the sperm extract.


CONCLUSIONS
Synthetic peptides of 80 kDa HSA are immunogenic and antibodies raised against these peptides recognize the native protein detected by ELISA, Western blot analysis. In addition, they possess sperm agglutinating activity. These findings suggest that they are promising candidates in the development of immunocontraceptives.
Vernekar
Vandana J
VJ
National Institute for Research, in Reproductive Health, J. M. Street, Parel, Mumbai, India.
Bandivdekar
A H
AH
Raghavan
Vijaya P
VP
Kamada
M
M
Koide
S S
SS
D000372:Agglutination Tests; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D000918:Antibody Specificity / Q000276:immunology; D000937:Antigen-Antibody Reactions / Q000276:immunology; D000941:Antigens / Q000737:chemistry / Q000276:immunology*; D003269:Contraception, Immunologic; D004797:Enzyme-Linked Immunosorbent Assay; D006433:Hemocyanins / Q000276:immunology; D006801:Humans; D007114:Immunization; D008297:Male; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D011817:Rabbits; D013094:Spermatozoa / Q000276:immunology*
D000906:Antibodies; D000941:Antigens; D010455:Peptides; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin