Juan G. RipollNicolás A. GiraldoNatalia I. BolañosAdriana CuéllarNubia RoaFernando RosasVíctor Manuel VelascoZulma M. CucunubáConcepción J. PuertaJohn Mario González2026-03-222026-03-22201410.4049/jimmunol.192.supp.67.1https://doi.org/10.4049/jimmunol.192.supp.67.1https://andeanlibrary.org/handle/123456789/60411Abstract Background: Chagas disease is a chronic parasitic infection whose pathogenesis is related to parasite persistence and a dysfunctional cellular immune response. There is no method to determine the pool of antigen-specific T cells in Trypanosoma cruzi-infected patients. Thus, the goal of this work was to determine the fraction of antigen-specific T cells by measuring the expression of CD154 and membrane TNFα on the bulk of parasite-stimulated PBMCs. Methodology: Twenty-one chagasic patients (CP), 11 healthy controls (HC) and 5 non-chagasic individuals with chronic heart disease (NCC) were analyzed. PBMCs were incubated in the presence of anti-CD28, anti-CD49d, anti-TNFα and TACE inhibitor as follows: non stimulus, T. cruzi-lysate or SEB. Later, the samples were stained for CD3, CD4, CD8 and CD154 and analyzed by flow cytometry. Results: Symptomatic (3.07%, SD ±1.98) and asymptomatic CP (1.24 ±0.59) displayed higher frequencies of CD4+/CD154+ cells after incubation with T. cruzi-lysate in comparison with NCC (0.084 ±0.03) and HC (0.11 ±0.08) donors P<0.0001. Likewise, the frequency of CD8+ cells expressing membrane TNFα with parasite stimulus was higher in symptomatic (3.76% ±1.43) and asymptomatic patients (2.22 ±0.53) compared with NCC (1.28 ±0.45) and HC (0.57 ±0.41) donors P<0.0001. Conclusions: These results suggest that membrane TNFα and CD154 after stimulation with T. cruzi-lysate could be useful to determine the fraction of parasite-specific T cells.enCD154Trypanosoma cruziAntigenImmune systemTumor necrosis factor alphaImmunologyChagas diseaseCD28CD8Peripheral blood mononuclear cellarticle