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.
Highlights
Chagas disease (CD) is a potentially life-threatening zoonosis caused by the hemoprotozoan parasite Trypanosoma cruzi, which may be transmitted by contact with feces/urine of infected blood-sucking triatomine bugs, consumption of contaminated food or beverages, blood transfusion, tissue and organ transplantation, from mother-to-child during pregnancy, and laboratory accidents [1]
After cell-disruption and centrifugation, recombinant proteins were recovered from the supernatant and the purity was estimated to be over 95% (Fig 1)
We aimed to produce and validate four chimeric recombinant antigenic proteins for the precise detection of anti-T. cruzi antibodies in sera from individuals infected with T. cruzi
Summary
Chagas disease (CD) is a potentially life-threatening zoonosis caused by the hemoprotozoan parasite Trypanosoma cruzi, which may be transmitted by contact with feces/urine of infected blood-sucking triatomine bugs, consumption of contaminated food or beverages, blood transfusion, tissue and organ transplantation, from mother-to-child during pregnancy, and laboratory accidents [1]. Diagnostic methods for detecting T. cruzi infection need to consider both clinical symptoms as well as the stage of illness. In the initial or so-called “acute” phase, the parasite can be detected in blood smears owing to high levels of parasitemia. At this stage, PCR-based methods can be adopted as efficient diagnostic tools, because specific antibodies can only be found several weeks after infection. Parasitemia is low but high levels of specific anti-T. cruzi antibodies (IgG) can be found in patient’s blood [5]. CD diagnosis requires the use of antigen-antibody detection methods, which are carried out by immunological techniques
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