Please use this identifier to cite or link to this item: https://www.arca.fiocruz.br/handle/icict/11536
Title: Chagas disease-specific antigens: characterization of epitopes in CRA/FRA by synthetic peptide mapping and evaluation by ELISA-peptide assay
Authors: Bottino, Carolina G.
Gomes, Luciano P.
Pereira, José B.
Coura, José Rodrigues
Provance Jr., David William
De Simone, Salvatore Giovanni
Affilliation: Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Bioquímica de Proteínas e Peptídeos. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Bioquímica de Proteínas e Peptídeos. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Doenças Parasitárias. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Doenças Parasitárias. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Centro de Desenvolvimento Tecnológico em Saúde (CDTS). Instituto Nacional de Ciência e Tecnologia para Inovação em Doenças Negligenciadas. Rio de Janeiro, RJ, Brasil
Fundação Oswaldo Cruz. Centro de Desenvolvimento Tecnológico em Saúde (CDTS). Instituto Nacional de Ciência e Tecnologia para Inovação em Doenças Negligenciadas. Rio de Janeiro, RJ, Brasil / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Bioquímica de Proteínas e Peptídeos. Rio de Janeiro, RJ, Brasil / Universidade Federal Fluminense. Instituto de Biologia. Departamento de Biologia Celular e Molecular. Niterói, RJ, Brasil.
Abstract: Background: The identification of epitopes in proteins recognized by medically relevant antibodies is useful for the development of peptide-based diagnostics and vaccines. In this study, epitopes in the cytoplasmic repetitive antigen (CRA) and flagellar repetitive antigen (FRA) proteins from Trypanosoma cruzi were identified using synthetic peptide techniques and pooled sera from Chagasic patients. The epitopes were further assayed with an ELISA assay based on synthetic peptides. Methods: Twenty-two overlapping synthetic peptides representing the coding sequence of the T. cruzi CRA and FRA proteins were assessed by a Spot-synthesis array analysis using sera donated by patients with Chagas disease. Shorter peptides were selected that represented the determined epitopes and synthesized by solid phase synthesis to evaluate the patterns of cross-reactivities and discrimination through an ELISA-diagnostic assay. Results: The peptide Spot-synthesis array successfully identified two IgG antigenic determinants in the CRA protein and four in FRA. Bioinformatics suggested that the CRA antigens were unique to T. cruzi while the FRA antigen showed similarity with sequences present within various proteins from Leishmania sp. Subsequently, shorter peptides representing the CRA-1, CRA-2 and FRA-1 epitopes were synthesized by solid phase synthesis and assayed by an ELISA-diagnostic assay. The CRA antigens gave a high discrimination between Chagasic, Leishmaniasis and T. cruzi-uninfected serum. A sensitivity and specificity of 100% was calculated for CRA. While the FRA antigen showed a slightly lower sensitivity (91.6%), its specificity was only 60%. Conclusions: The epitopes recognized by human anti-T. cruzi antibodies have been precisely located in two biomarkers of T. cruzi, CRA and FRA. The results from screening a panel of patient sera through an ELISA assay based on peptides representing these epitopes strongly suggest that the sequences from CRA would be useful for the development of diagnostic reagents that could improve upon the sensitivity and specificity of currently available diagnostic tests. Overall, the results provide further evidence of the usefulness of identifying specific linear B-cell epitopes for improving diagnostic tools.
Keywords: Chagas Disease
Trypanosoma cruzi
Cytoplasmic repetitive antigen
Flagellar repetitive antigen
Epitopes
Spot-synthesis
Peptide-ELISA
DeCS: Doença de Chagas
Trypanosoma cruzi
Epitopos
Ensaio de Imunoadsorção Enzimática
Peptídeos
Issue Date: 2013
Publisher: BioMed Central
Citation: BOTTINO, Carolina G.; et al. Chagas disease-specific antigens: characterization of epitopes in CRA/FRA by synthetic peptide mapping and evaluation by ELISA-peptide assay. BMC Infectious Diseases, v.13:568, 10p, 2013.
DOI: 10.1186/1471-2334-13-568
Copyright: open access
Appears in Collections:CDTS - Artigos de Periódicos
IOC - Artigos de Periódicos

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