Immunogenicity of malaria transmission-blocking vaccine candidate, y230.CA14 following crosslinking in the presence of tetanus toroid

Proteolytically processed 310 kDa form of Plasmodium falciparum gamete surf ace antigen, Pfs230, is the target of malaria transmission-blocking monoclo nal antibodies. To design a recombinant malaria transmission-blocking subun it vaccine, the amino terminus of the 310 kDa surface-exposed form of Pfs23 0 was mapped to amino acids (aa) 522 and 584 using a series of peptides and recombinant proteins encoding distinct regions of Pfs230. Antiserum genera ted against an Escherichia coli-produced recombinant protein, spanning the Pfs230 processing site and extending into the cysteine domains, r230/MBP.C (aa 443-1132), reduced parasite infectivity by 71.2-89.8%. To determine if the region spanning the cleavage site blocked malaria transmission when pro duced as a secreted protein by Saccharomyces cerevisiae, y230.CA14 (aa 467- 584) was generated, purified emulsified in adjuvant and used to vaccinate m ice. In contrast to E. coli-produced r230/MBP.C, the immune response genera ted against y230.CA14 was very weak. To enhance the response, y230.CA14 was mixed with tetanus toroid, chemically cross-linked, repurifed, and its imm unogenicty compared with unconjugated y230.CA14. Conjugated-y230. CA14/TT r equired fewer booster injections to induce an immune response against Pfs23 0 and the antibodies generated reacted with the surface of intact gametes a nd immunoprecipitated radiolabelled Pfs230 extracted from I-125 surface-lab elled gametes to a greater extent. After seven injections, all y230.CA14 va ccinated mice developed anti-Pfs230 antibodies and the isotype profile was the same. in addition to enhancing the initial immune response generated ag ainst y230. CA14, conjugation focuses the immune response toward epitopes w ithin the region of Pfs230 present on the surface of the gamete.