Evaluating Immune Cell Responses in Mice

To determine immune cell responses, the levels of T and B cell populations from blood, spleen, and ILN of mice were investigated by flow cytometry as described previously [8 (link)]. CD4⁺ and CD8⁺ T cells from the blood were detected after 2nd boost immunization and on day 6 after challenge infection. The levels of CD4⁺, CD8⁺ T cells, and germinal center B cells from inguinal lymph node (ILN) cells were detected 6 days after challenge infection. Immune cells ( $1\times {10}^6$ cells in each tube) resuspended in PBS were incubated with stimulants at a concentration of 0.05 μg/100 μL of P. berghei antigen for 2 h at 37 °C or left unstimulated. The cells were incubated with the surface marker antibodies (CD3e-PE-Cy5, CD4-FITC, CD8a-PE, B220-FITC, GL7-PE; BD Biosciences, CA, USA) at 4 °C for 30 min. Stained cells were acquired using a BD Accuri C6 Flow Cytometer (BD Biosciences, CA, USA) and data were analyzed using C6 Analysis software (BD Biosciences, CA, USA).