FIG 2.

Animal experimental design (Experiment 1) and intracellular production of IL-4, IFN-γ, and TNF-α in CD4 and CD8 T cells. An illustration of the animal experiment (Experiment 1) is shown (A). C57BL/6 mice were orally administered the appropriate material three times at three-week intervals. The intracellular cytokine (ICS) assay was performed at 10 days post-second immunization (10 dp2i) and 10 days post-third immunization (10 dp3i) by fluorescence-activated cell sorting (FACS). An ELISpot and an ELISA were also conducted at 10 dp3i. The remaining mice were challenged with 0.5 LD₅₀ H1N1 virus 9 weeks after the first immunization, and their body weights were monitored for 2 weeks. The single-cell suspensions from the spleen samples were subjected to FACS to determine the intracellular cytokine (IL-4, IFN-γ, and TNF-a) production in both the CD4⁺ cells (B) and the CD8⁺ (C) T cells at 10 dp2i (n = 3) and 10 dp3i (D and E) (n = 4; *, P < 0.05; **, P < 0.01; ***, P < 0.001).