Performance Assessment of Four Chimeric Trypanosoma cruzi Antigens Based on Antigen-Antibody Detection for Diagnosis of Chronic Chagas Disease

Abstract

The performance of serologic tests in chronic Chagas disease diagnosis largely depends on the type and quality of the antigen preparations that are used for detection of anti-Trypanosoma cruzi antibodies. Whole-cell T. cruzi extracts or recombinant proteins have shown variation in the performance and cross-reactivity. Synthetic chimeric proteins comprising fragments of repetitive amino acids of several different proteins have been shown to improve assay performances to detect Chagasic infections. Here, we describe the production of four chimeric T. cruzi proteins and the assessment of their performance for diagnostic purposes. Circular Dichroism spectra indicated the absence of well-defined secondary structures, while polydispersity evaluated by Dynamic Light Scattering revealed only minor aggregates in 50 mM carbonate-bicarbonate (pH 9.6), demonstrating that it is an appropriate buffering system for sensitizing microplates. Serum samples from T. cruzi-infected and non-infected individuals were used to assess the performance of these antigens for detecting antibodies against T. cruzi, using both enzyme-linked immunosorbent assay and a liquid bead array platform. Performance parameters (AUC, sensitivity, specificity, accuracy and J index) showed high diagnostic accuracy for all chimeric proteins for detection of specific anti-T. cruzi antibodies and differentiated seropositive individuals from those who were seronegative. Our data suggest that these four chimeric proteins are eligible for phase II studies.

Accuracy of chimeric proteins in the serological diagnosis of chronic chagas disease – a Phase II study

“…A study in Argentina, for instance, performed in a co-endemic area for leishmaniasis and chronic CD, reported 81.8% (9/11) cross-reactivity using conventional ELISA [51]. A more recent study confirmed this elevated cross-reaction rate and verified a relatively large number of inconclusive results (2.9–8.6%) using conventional and non-conventional tests [9]. Contrary to these data, IBMP-8.1 chimera cross-reacted with two samples and IBMP-8.3 and -8.4 with one sample, demonstrating very little positivity in 153 sample of Leishmania spp.…”

Performance Assessment of a Trypanosoma cruzi Chimeric Antigen in Multiplex Liquid Microarray Assays

“…Therefore, chimeric proteins have been proposed to improve the assay's accuracy to diagnose CD. Recently, a phase I study was performed with 4 chimeric proteins from the Instituto de Biologia Molecular do Paraná (IBMP-8.1, 8.2, 8.3, and 8.4) to detect specific anti-T. cruzi antibodies using both ELISA and LMA (15), demonstrating that each antigen accurately discriminated CD-positive from CD-negative samples. In addition, no significant differences were observed with respect to the diagnostic performances of the ELISA and LMA test methods.…”

Electrochemical Biosensing for the Diagnosis of Viral Infections and Tropical Diseases

“…[90] Various strategies for diagnosis of Chagas disease and trypanosomiasis using electrochemical biosensors have been reported. [61,62,[91][92][93][94][95][96][97] Diniz et al [91] adsorbed a polypeptide chain formed by recombinant antigens, cytoplasmic repetitive antigen and flagellar repetitive antigen of Trypanosoma cruzi, onto an oxidized gold electrode, and the interaction with sera from chronic chagasic patients was monitored by electrochemical impedance spectroscopy. More recently, Ferreira et al [62] used T.cruzi proteins from epimastigote membranes for the construction of an amperometric immunosensor for serological diagnosis of Chagas' disease.…”

“…An immunosensor for anti-T.cruzi was developed by immobilization the antigen and EIS measurement of antibody interaction. [95] Qualitative diagnosis of Chagas' disease for verifying the presence or the absence of T cruzi using a simple configuration was also proposed by Santos et al [96,97] Direct detection of the immunoreaction was performed by EIS after immobilization of the antigen onto a gold electrode modified with a MPA selfassembled monolayer (SAM). Recently, a sandwich immunosensor for anti-IgM T. cruzi-specific antibodies, which are a biomarker for congenital Chagas disease was also reported.…”