DETECTION OF CRUZIPAIN, THE MAJOR CYSTEINE PROTEINASE FROM TRYPANOSOMA-CRUZI AND ITS C-TERMINAL EXTENSION IN BIOLOGICAL-FLUIDS DURING EXPERIMENTAL-INFECTION IN MICE

Monoclonal antibodies (MoAbs) specific for unique epitopes of the cata lytic domain of cruzipain (Crz) were used to develop a two-site sandwi ch ELISA specific for native Crz. In addition, the authors developed a sandwich ELISA that allowed the detection of the protease C-terminal domain (CT) using a combination of a MoAb specific For the CT and rabb it anti-Crz IgGs. Both assays were sensitive with detection limits of 2 ng/ml and 0.7 ng/ml, respectively. The assays were assessed for appl icability in detection of antigens in serum and urine from experimenta lly infected BALB/c mice. The antigens were already detectable in seru m by the third week after infection, reached their peak by week four, and decreased during the chronic phase of the infection. Throughout th e infection the relative amount of CT detected was several-fold higher than that of native Crz, and the data demonstrate that the cT exposes highly immunogenic epitopes that are absent in native Crz. Since thes e observations have a potential application in diagnosis, the authors analysed the degree of cross-reactivity with antigens from T. rangeli, T. brucei, Leishmania mexicana and L, panamensis, and determined that the assays were highly specific, Measurable amounts of the CT were al so recorded in urine samples.