Mouse Airway Epithelial Cell Isolation and Differentiation

The preparation of primary mouse airway epithelial cells (AECs) was performed as previously described (24 (link)). Briefly, cells were isolated from the tracheae of mice with the indicated genotypes by enzymatic treatment and seeded onto a 0.4-μm pore size clear polyester membrane (Corning) coated with a collagen solution. At confluency, the medium was removed from the upper chamber to establish an air-liquid interface. Fully differentiated, 7-10-day-old post-air-liquid interface cultures were used routinely for experiments. For IFN treatment, cells were treated for 4 h from the basolateral side with indicated concentrations of either IFN-α_B/D or recombinant mouse IFN-λ2 and then processed for RNA isolation and subsequent RT-qPCR analysis. For time course analysis, AECs were treated for 1 h with 10 ng/ml of either IFN-α_B/D or recombinant mouse IFN-λ2 and processed for RNA isolation and subsequent RT-qPCR analysis.