Fresh tissue specimens were collected using standard institutional procedures and stored in MACS Tissue Storage Solution (Miltenyi Biotec) at 4°C for up to 24 hours prior to processing.  Biopsy tissue was dissociated mechanically using a scalpel and enzymatically using the Miltenyi Biotec Human Tumor Dissociation Kit (Order no. 130-095-929) and a gentleMACS Dissociator 2.2.2. The dissociated sample was cryopreserved in 90%FBS 10%DMSO and stored in liquid nitrogen prior to cell isolation. For CD45+ cell isolation, samples were thawed and isolated using the Miltenyi Biotec Human CD45 (TIL) MicroBeads using magnetic separation with MS columns (Order no. 130-118-780). All samples were depleted during sample preparation. Cell viability was determined using the Muse Count & Viability Reagent (Millipore). Samples with greater than 60% viability were processed for scRNA-seq using the 10X Genomics 3' Gene Expression v3.1 kit (10X Genomics Protocol #CG000315). For each sample, 10,000 cells were targeted. Libraries were sequenced on an Illumina NovaSeq6000.