Tissues were processed in RPMI supplemented with 10% fetal bovine serum, 83 μg/ml gentamicin, and 1% GlutaMAX, henceforth referred to as RPMI complete media, digested in RPMI containing collagenase and DNase, and washed with RPMI complete media. After passage through a 40μm strainer, a single cell suspension was achieved. Dead cells were removed using the Dead Cell Removal Kit (STEMCELL Technologies). CD45+ cells were then isolated with the CD45+ isolation kit (STEMCELL Technologies). To increase the recovery of Kupffer cells, the flow through from the CD45+ isolation kit was then layered over Ficoll Plaque Plus (Millipore Sigma) and spun at 2200 rpm for 20 minutes. Cells from the buffy coat were then combined with the CD45+ cells and passed through a 35μm strainer to ensure a single cell suspension. Next-generation sequencing libraries were prepared using the 10x Genomics Chromium Single Cell 3' Reagent kit v3.1 per manufacturer's instructions. Libraries were uniquely indexed using the Chromium dual Index Kit, pooled, and sequenced on an Illumina NovaSeq 6000 sequencer in a paired-end, dual indexing run. Sequencing for each library targeted 20,000 mean reads per cell.