Immunofluorescence Staining of HBEC

Immunofluorescence staining of HBEC was performed as previously described^{20 ,25 (link)}. ALI cultures were fixed with a solution of 50%/50% methanol/acetone for 2 min at – 20 °C followed by three washes with PBS. Fixed cultures were incubated with anti-MUC5AC antibody (MA1-38223; Thermo Fisher Scientific) at 0.4 µg/mL and anti-MUC5B antibody (#37-7400; Thermo Fisher Scientific) at 2 µg/mL overnight at 4 °C. Cultures were then incubated with Alexa Fluor 555 donkey anti-mouse IgG (#A31570; Thermo Fisher Scientific) at 1 µg/mL and Alexa Fluor 488 donkey anti-rabbit IgG (#A21206; Thermo Fisher Scientific) at 1 µg/mL for 1 h at room temperature. Anti-acteylated α-tubulin antibody (#T6793; MilliporeSigma) was used at 1:1000 dilution overnight at 4 °C. Hoechst 33258 (#H3569; Thermo Fisher Scientific) was used at 2 µg/mL for 10 min at room temperature.