Technology type 'Immunostaining'

A549 cells were seeded in 96-well plates at a density of 2 × 104 cells per well. The following day, virus stocks were serially diluted in DMEM containing 2% FBS (2% DMEM), and cells were inoculated with dilutions in triplicate at 37°C for 2 h. After the incubation, cells were overlaid with DMEM supplemented with 1% carboxymethylcellulose (Fisher Scientific), 2% FBS, 10 mM HEPES, and 1× penicillin/streptomycin. At 48 hpi, the methylcellulose overlay and medium were removed, and cells were fixed ...

For immunoblot analysis, protein lysates were incubated with 4× Laemmli buffer (Bio-Rad) containing 10% β-mercaptoethanol (Thermo Fisher-Life Technologies) at 95°C for 3 min. Approximately 7 to 10 μg of total protein was loaded per lane onto 4 to 20% Criterion TGX gradient gels (Bio-Rad) and electrophoresed at ~96 V in 1× sodium dodecyl sulfate (SDS) running buffer (25 mM Tris, 192 mM glycine, 0.1% SDS). Proteins were then transferred onto nitrocellulose membranes (Fisher Scientific) at 90 V for ...