Development and evaluation of a SPR-based immunosensor for detection of anti-Trypanosoma cruzi antibodies in human serum

Abstract

Protozoan Trypanosoma cruzi is the etiological agent of Chagas disease, which needs urgent progress in its immunological diagnosis. Surface plasmon resonance (SPR) is a promising technique for the development of immunosensors with biomedical applications. In this work, a SPR-based immunosensor has been developed by the first time for real time and label free immunoassay for detection of anti-T. cruzi antibodies in serum samples. T. cruzi antigen was successfully immobilized on a SPR sensor chip via activated mixed self-assembled monolayer (SAM) of 3-mercaptopropionic acid (3-MPA) and 11-mercaptoundecanoic acid (11-MUA), by amide coupling. After the sensor construction, a pool of human sera infected with T. cruzi was added to its surface and the antibodies were detected in sera diluted up to 1280 times, indicating excellent sensitivity of the technique for detection of antigen–antibody interaction. The addition of a pool of negative human serum at dilutions lower than 1:160 to the sensor surface was accompanied by a null or very low response. Then, the following operational parameters of the immunoassay were optimized and defined: time of immobilization and antigen concentration at 20min and 30mgmL−1, serum dilution at 1:320, preventing of nonspecific bindings with solution of BSA 1.0% and surface regeneration by injection of SDS 1.0%. The immunoassay, here termed SPRCruzi, showed high capability in the discrimination of positive and negative sera, including those infected with other pathogens usually sources of false positives results in conventional serodiagnosis. Therefore, the proposed immunosensor was successfully developed and the immunoassay allowed a simple, effective, faster and specific detection of anti-T. cruzi antibodies, which represents an encouraging field for the progress of the diagnosis of Chagas disease.