Mouse anti epcam antibodies

Mouse AEC cultures were prepared from female C57BL/6 mice as previously described22 (link), with minor modifications. Briefly, lungs from mice were digested in 1.5 mg/ml Pronase (Roche, USA), 0.1 mg/ml DNase I (Sigma-Aldrich, USA) for 1 hr at 37°C in 5% CO₂. Single cell suspensions were incubated with purified rat anti-mouse CD45 antibody (BD Biosciences, USA) and epithelial cells negatively enriched with BioMag goat anti-rat Ig-coupled magnetic beads (Qiagen, USA). Cells were cultured on collagen-coated (MP Biomedicals, USA) plates in DMEM/F-12 media (Gibco, USA) supplemented with 10% FCS, 2 mM L-glutamine, 100 U/ml penicillin, 100 μg/ml streptomycin and 5 μg/ml insulin (Sigma-Aldrich). To confirm cell purity, monolayers were detached with 3 mM EDTA and cells stained with mouse anti-EpCAM antibodies (BioLegend, USA) and examined by flow cytometry. Cell monolayers were negative for the presence of fibroblasts (anti-mouse vimentin (Santa Cruz, USA)). Monolayers were infected with the HKx31 strain of IAV (H3N2, MOI 1) in serum free DMEM/F-12 for 1 hr and then washed and incubated in DMEM/F-12 supplemented with 2% FCS and 4 μg/ml trypsin for 24 or 48 hrs.