Klrg1 clone 2f1 klrg1

Tumor tissue was mechanically dissociated and digested in 0.7 mg/mL Collagenase D (Roche) and 3 mg/mL DNase I (Roche) for 30–45 min to obtain single-cell suspension. For experiments looking at tumor-infiltrating leukocytes (TIL), TIL were isolated using Percoll gradient prior to staining. For experiments looking at B16RFP⁺ tumor cells, cells were stained immediately after digestion for 20 min. Cells were blocked with anti-CD16/32 (clone 93; Biolegend) prior to surface staining. Antibodies against the following antigens were used: CD11a (clone M17/4; ThermoFisher), CD8a (clone 53-6.7; BD Biosciences or Biolegend or eBioscience), CD45 (clone 30-F11; Invitrogen or eBioscience), CD45.1 (clone A20; Biolegend), CD3 (clone eBio500A2; eBioscience), CD45.2 (clone 104; eBioscience), CD90.1 (clone OX-7; BD Biosciences), CD127 (clone A7R34; eBioscience or clone SB/199; Biolegend), KLRG1 (clone 2F1/KLRG1; Biolegend or eBioscience), PD-1 (clone RMP1-30; eBioscience), NKG2A/C/E (clone 20d5; eBioscience), LAG3 (clone C9B7W; Biolegend), CD44 (clone IM7; eBioscience or Biolegend or BD Biosciences), Ly6C (clone HK1.4; Biolegend), Ly6G (clone 1A8; Biolegend), CD11b (M1/70; eBioscience or Biolegend), Qa-1^b (clone 6A8.6F10.1A6; Miltenyi Biotec), and PD-L1 (10F.9G2; Biolegend).

The following antibodies were purchased from BioLegend (San Diego, CA): CD8 (clone 53–6.7)-PerCP/Cy5.5 and -PE, CD4 (clone RM4-5)-PerCP/Cy5.5, CD27 (clone LG.3A10)-PE/Cy7, CD43 (clone 1B11)-APC, CD62L (clone MEL-14)-PE, CD90.2 (Thy1.2) (clone 53–2.1)-FITC, CD127 (clone A7R34)-Brilliant Violet 421, KLRG1 (clone 2F1/KLRG1)-APC, IFN-γ (clone XMG1.2)-FITC, TNF (clone MP6-XT22)-PE, IL-2 (clone JES6-5H4)-Brilliant Violet 421, Rat IgG2a (clone RTK2758)-PE, -APC, -Brilliant Violet 421, Rat IgG2b (clone RTK4530)-Brilliant Violet 421, Rat IgG1 (clone RTK2071)-PE, Syrian Hamster IgG (clone SHG-1)-APC, Armenian Hamster (clone HTK888)-PE/Cy7.