Protective, lytic antibodies are believed to be correlated with active Trypanosoma cruzi infection. In patients with chronic infection, antibodies lysing trypomastigote forms recognize chiefly alpha-galactosyl structures at the parasite surface. The target molecules on cell-derived trypomastigotes that react with anti-alpha-galactosyl antibodies (anti-Gal) from patients with chronic Chagas disease were investigated. Glycoconjugates were isolated from trypomastigotes and shown to absorb purified Chagasic (Ch) anti-Gel effectively as well as lytic antibodies from Ch sera. Active fractions were F2 (74 kD and 95.6 kD) and F3 (120-200 kD). A differential reactivity with antibodies from untreated Ch patients (trypanolytic) and from treated, presumably cured, individuals (not trypanolytic) was evident using F2 and F3 antigenic fractions. No cross-reactivity with heterologous sera (other infections) was observed. The F2 glycoconjugate (mostly 74 kD) can be used in the diagnosis of active Chagas infection, replacing the quantitative determination of complement-mediated lysis. With the present sample of patients' sera and normal human sera, it showed 100% sensitivity and specificity.