Characterizing T and B Cell Responses

Splenocytes and MLN cells were prepared for flow cytometric analysis as previously described (Kang et al., 2020a (link)). Single cell suspensions of splenocytes (1 x 10⁶ cells/mouse) and MLN cells (1 x 10⁵ cells/mouse) were stimulated ex vivo with T. gondii ME49 antigen (2 μg/mL) at 37°C with 5% CO₂ for 2 hours. After antigen stimulation, cells were stained with the following fluorescent-conjugated antibodies purchased from BD Biosciences (Franklin Lakes, NJ, USA) and Invitrogen (Waltham, MA, USA) for CD4⁺ T cell, CD8⁺ T cell, and germinal center B (GC B) cell detection: CD3 (PE-Cy7), CD4 (FITC), CD8 (PE), GL7 (PE), and B220 (FITC). All staining procedures for flow cytometry were performed according to the manufacturer’s protocol. Stained cells were acquired using the Accuri C6 flow cytometer and analyzed with the C6 Accuri software (BD Biosciences, Franklin Lakes, NJ, USA).