MDCK cells were cultured in E10 medium comprising Dulbecco's modified Eagle medium (DMEM, Invitrogen, Carlsbad, CA) supplemented with 10% fetal bovine serum (FBS; JRH Biosciences, Lenexa, KS, USA), 100 U/mL penicillin, 100 μg/mL streptomycin (Sigma-Aldrich, St Louis, MO, USA), 2 mM L-glutamine (Gibco BRL, Gaithersburg, MD, USA), and 0.1 mM nonessential amino acid mixture (Gibco). The cells were maintained in a 37 °C incubator with 5% CO2. The influenza virus A/WSN/33 (WSN) obtained from the American Type Culture Collection was propagated in MDCK cells. Cells were incubated with the influenza virus at a multiplicity of infection (MOI) of 0.5 for 1 h at room temperature and cultured for 24 or 48 h at 37 °C in fresh DMEM. Viral titers in supernatants were determined using MDCK cells by the 50% tissue culture infectious dose (TCID50) assay .