Evaluation of in-house ELISA using Trypanosoma cruzi lysate and recombinant antigens for diagnosis of Chagas disease and discrimination of its clinical forms

Abstract

The aim of this work was to investigate the potential usefulness of Trypanosoma cruzi lysate, recombinant protein JL7, and peptides P013, R13, JL18, JL19, and P0β as serological markers for human Chagas disease. We analyzed 228 sera from Brazilian Chagas disease patients classified into four clinical groups and 108 from non-chagasic patients. We defined the diagnostic sensitivity, specificity, and Kappa index measured by enzyme-linked immunosorbent assay (ELISA). As previously described, the highest values of diagnostic parameters were achieved for T. cruzi lysate and JL7; peptide P013 showed high specificity but low sensitivity. The other peptides resulted in lower sensitivity and specificity in our ELISA than T. cruzi lysate and JL7 protein. Antibodies against JL7 protein were mainly detected in sera from patients with severe chagasic cardiomyopathy, compared with those from the indeterminate form, whereas peptides failed to discriminate between the clinical forms of the disease.

Figure 1.

Performance of antigens for prognosis of Chagas disease. Enzyme-linked immunosorbent assay (ELISA) plates were coated with 50 ng protein/well of Trypanosoma cruzi epimastigote lysate, 2 μg/well of recombinant protein JL7 or 2 μM of peptides coupled to bovine serum albumin (BSA) in 50 μL buffer carbonate. Serum samples were diluted 1/400 and tested in duplicate. Sera from six healthy individuals were loaded on each plate to determine cut-off value, as the optical density (OD) mean value plus three standard deviations. Results were expressed as ratio calculated as (OD value of each serum samples/cut-off value) ±SE. IC = indeterminate form of Chagas disease; DC = digestive form of Chagas disease; MCC = mild chagasic cardiomyopathy; SCC = severe chagasic cardiomyopathy.