ACTIVATION-INDUCED MARKER ASSAY, AN ALTERNATIVE STRATEGY TO STUDY TRYPANOSOMA CRUZI SPECIFIC CD8+ T CELLS

LUCIA BISCARI; CINTIA KAUFMAN; CECILIA FARRÉ; VICTORIA HUHN; ANA ROSA PÉREZ; ANDRÉS ALLOATTI

Virtual

Congreso; Reunión Anual de Sociedades de Biociencias 2021; 2021

Sociedad Argentina de Inmunología, Sociedad Argentina de Investigación Clínica, Asociación Argentina de Farmacología Experimental, Asociación Argentina de Nanomedicinas

In the investigation of the immune response induced by vaccines or generated against infectious diseases, the determination of the specific CD8 response is of particular importance. Although there are various techniques for this purpose, such as the use of specific tetramers or ELISPOT, the activation ? induced marker (AIM) technique emerges as a simpler experimental approach. In this work, we tested and optimized AIM to analyze the response of CD8+ T cells specific for Trypanosoma cruzi antigens, generated after infection of C57BL/6 mice with the Tulahuén strain of T. cruzi. Animals were infected intraperitoneally with 5000 T. cruzi trypomastigotes. At 12 days post infection, at the peak of the CD8+ T response, the animals were sacrificed and spleens and lymph nodes were harvested. Splenocytes and lymph node cells were incubated for 15 h with different concentrations of a particular peptide (TsKb20, derived from the Transialidase protein or PAR4, derived from the PAR4 protein of the flagellum, from T. cruzi). Cells incubated with Concanavalin A were used as a positive control, and non-restimulated cells were used as a negative control. The specific CD8 response was determined by flow cytometry evaluating the activation markers CD25+ and CD69+ in the population of CD8+ T cells. Through a two-way ANOVA analysis, it was found that the specific CD8+ T cell response for TsKb20 in infected animals, measured after restimulation with 50 ug/mL of the TsKb20 peptide, was significantly higher than the response measured in uninfected animals. No difference was evidenced in the specific CD8 response for PAR4. These results suggest that the AIM technique could be used to determine anti-T. cruzi specific CD8 response.