Tandem use of PCR and synthetic peptides to map helper T-cell epitopes on 27-kDa sexual stage antigen of Plasmodium falciparum.

Abstract

Monoclonal antibodies recognizing two overlapping linear epitopes (amino acid residues 10 to 25) on the 27-kDa sexual stage antigen of Plasmodium falciparum (Pfg 27) effectively reduce the infectivity of the parasites to mosquitoes. Although malaria transmission-blocking immunity is largely antibody-mediated, T cells play critical roles in the regulation of antibody-secreting B cells. In order to facilitate the development of a malaria transmission-blocking subunit vaccine, studies were undertaken to map epitopes on Pfg 27 recognized by T-helper lymphocytes. Pfg27-specific T-cell hybridoma clones were produced from rPfg27-immunized BALB/c (H-2d) and C57BL/6 (H-2b) mice, and used in studies to map antigenic determinants using PCR-generated Pfg27 gene fragments expressed in E. coli and synthetic peptides based on the Pfg27 sequence. We identified and mapped five distinct T-cell epitopes that are recognized by major histocompatibility complex (MHC) class II-restricted T-cell hybridoma clones. A single peptide (21 residues) was shown to contain two tandem or partially overlapping epitopes recognized by T-cell hybridomas in the context of I-Ad and I-Ab, respectively. Synthetic peptides representing epitopes recognized by T-cell hybridoma clones elicited strong IgG responses in immunized mice, suggesting that T-cells of the helper phenotype were stimulated in vivo by these peptides. These studies represent the first detailed T-cell epitope analysis of a malaria sexual-stage antigen.