MDCK and Calu-3 Cell Culture Protocols

Madin–Darby Canine Kidney (MDCK CCL-34) cells (ATCC, Manassas, VA, USA) were cultured at 37 °C and 5% CO₂ in Dulbecco’s Modified Eagle Medium (DMEM, high glucose pyruvate; Gibco, Waltham, MA, USA). DMEM was supplemented with 10% foetal bovine serum (FBS, Bovogen Biologicals, Australia), 1× GlutaMAX (Gibco, USA), 1× MEM non-essential amino acid solution (Gibco, USA), 0.05% sodium bicarbonate (Gibco, USA), 20 μM HEPES (Gibco, USA), and 100 U/mL penicillin-streptomycin solution (Gibco, USA). Maintenance medium (DMEM medium containing the above constituents excluding only serum) was used for virus dilutions. Maintenance medium supplemented with 2 µg/mL TPCK-treated trypsin (Infection medium; SAFC Biosciences, Lenexa, KS, USA) was used for virus infection protocols [3 (link)].
Calu-3 (HTB-55) cells (ATCC, USA) were cultured at 37 °C and 5% CO₂ in Eagle’s Minimum Essential Medium (EMEM; Gibco, USA), supplemented with 10% FBS, 50 U/mL penicillin-streptomycin solution, 1× MEM non-essential amino acid solution, and 1 mM sodium pyruvate (Gibco, USA). During infection, Calu-3 medium was supplemented as above, although modified to include 0.3% FBS and 2 µg/mL TPCK-treated trypsin.