Fig. 5

Flow cytometry of immune cells in peripheral blood of mice demonstrated a successful immune reconstitution. A Gating strategy of lymphocytes including T cells and B cells. Total leukocytes were gated with CD45⁺, in total lymphocytes were gated with CD3⁺. Then the CD4⁺ helper T cells, CD8⁺ cytotoxic T cells, and CD19⁺ B cells were then delineated in CD3⁺ lymphocytes, respectively. Next, CD4⁺ T cells were further divided into early activation (CD69⁺), middle activation (CD25⁺), late activation (HLA-DR⁺); exhausted (PD1⁺/CTLA-4⁺/TIM3^+/TIGIT⁺), as well as immunosuppressive Treg (FOXP3⁺CD25⁺) T cells. Similarly, we also applied the above gating strategy in CD8⁺T cells. In addition, we checked IFN-γ and Granzyme B, which were related to killing functions. B Gating strategy of myeloid cells involving macrophages and MDSCs. Among CD45⁺ leukocytes, overall myeloid lineage cells were delineated with CD11b⁺. Total macrophages were gated with CD68⁺; classically activated M1 macrophages were gated with CD86⁺; conditionally activated M2 macrophages were gated with CD206⁺; and immunosuppressive MDSCs were circled with HLA-DR-CD33⁺. C Comparison of the proportion of major cell components of tumor-infiltrating immune cells in each treatment group