Fig. 4: Evaluation of circulating T cells before and after neoadjuvant ICI treatment.

a, General gating strategy for PBMC samples. Left to right, a debris exclusion gate, a time gate to exclude electronic noise, a singlet gate to exclude doublets and a viability gate to exclude dead cells. T cells (CD3⁺ SSC-A-low) were gated for CD45RA⁻CD45RO⁺ effector and memory subsets: CD8⁺ T_EM (CD3⁺CD8⁺CD45RA⁻CD45RO⁺), CD4⁺ T_EM (CD3⁺CD4⁺FOXP3⁻CD45RA⁻CD45RO⁺) and T_reg (CD3⁺CD4⁺FOXP3⁺CD45RA^–CD45RO⁺) cells. b, CD8⁺ T_EM, CD4⁺ T_EM and T_reg subsets in the pretreatment (day −4, left) and posttreatment (day +12, right) samples were analyzed for (top to bottom): T cell-bound nivolumab (detected with anti-IgG4-PE), residual unoccupied PD-1 (direct PD-1 detection with anti-PD-1 (CD279) BV421), activation markers (MHC class II, OX40) and GzmB. Immune checkpoint (LAG3, CTLA-4, TIGIT) expression analysis was performed on pretreatment (day −9) samples. The numbers indicate the percentage of cells in the respective gates.

Source data