Single-cell suspensions of murine tumors and spleens were prepared as previously described.26 (link) Briefly, murine tumors were minced and digested in complete culture media containing 1 mg/mL of Collagenase IV (Roche) and 50 unit/mL DNase (Sigma) at 37°C with shaking every 10 min. After 30 min of incubation, 10 mL of fresh complete culture media was added and the samples were filtered through a 70 µM filter and centrifuged. Red blood cells were lysed on ice for 5 min with red blood cell lysis solution (BioLegend) followed by addition of complete media. Spleens and lymph nodes were mashed through a 70 µM filter and centrifuged. Red blood cells were lysed as described above, and the cells were washed with PBS, and counted.