RESUMO
To obtain antibodies suitable to be used in an antigen-capture assay, we have identified, synthesized, and evaluated a series of peptides from different Plasmodium falciparum excretory-secretory proteins: glutamate-rich protein (GLURP); histidine-rich protein 2; histidine-rich protein 3; Falciparum interspersed repeat antigen and, serine-rich antigen homologous. Conformational as well as antigenic predictions were performed using the ANTHEPROT package. Chemical synthesis was carried out by the multiple manual synthesis using the t-boc strategy. The peptides were used as antigens for the preparation of polyclonal antibodies in rabbits. Out of the 14 peptide constructs, eight by ELISA and, six by MABA elicited antibodies that showed correspondence between the predictive study and the immunogenicity obtained in rabbits. All antipeptide (GLURP, HRP2, and FIRA) antisera were found to bind to the corresponding synthetic sequence in an ELISA assay. The binding activity and specificity of antibodies were determined by Western blot with supernatant culture from P. falciparum. Anti-GLURP (IMT-94 and IMT-200) antisera bound to five molecules present in supernatant with molecular weight of 73, 82, 116, 124, and 128 kDa. Anti-HRP2 (IMT-192) antisera recognized a band of 58 kDa. In both cases, the specific molecules were inhibited by preincubation with the homologous peptide. Anti-HRP3, anti-FIRA neither anti-SERPH antisera showed reactivity. Anti-peptides HRP2 antibodies recognized the recombinant protein present in Parasight-F test. The same way, synthetic peptides from HRPII molecule were recognized by monoclonal antibody present in the Parasight-F assay. Our results confirm the potential value of synthetic peptides when inducing monospecific polyclonal antibodies for the development of diagnostic tests based on the capture of antigens.
