Lung Cell Dissociation and CD45+ Enrichment

Lung cells were dissociated as previously described (Sharma et al., 2022 (link)). The right lung was dissociated to a single cell suspension with Multi Tissue Dissociation Kit 2 (130–110–203, Miltenyi) according to the manufacturer’s protocol using the gentle MACS Dissociator (Miltenyi). The cell suspension was filtered through a 70 μm strainer, pelleted by centrifugation, and cleared of red blood cells with ACK Red Blood Cell Lysis buffer (BP10-548E, Lonza). Live cells were enumerated using trypan blue exclusion counting on a Countess Automated Cell Counter Hemocytometer C10227 (Invitrogen). Lung cells were then used as described below for Fluorescence-Activated Cell Sorting (FACS) analysis. Additionally, ten million lung cells per animal were used for CD45⁺ enrichment using the CD45 MicroBeads kit (cat# 130–109–682, Miltenyi) per manufacturer’s instructions.