Cell culture

The human lung adenocarcinoma A549 cells, acquired from Chinese Academy of Sciences, Shanghai Institute Cell Bank, were stored in a RPMI 1640 culture medium (Gibco Company, Grand Island, NY, U.S.A.), containing 10% FBS for incubation at 37°C in a 5% CO₂ incubator. Once the cells reached a confluence of approximately 80%, they were passaged. Following the removal of waste liquid, cells were washed twice by phosphate-buffered solution (PBS) and treated with 0.25% trypsin. Next, cells were made into suspension in RPMI 1640 culture medium containing 10% FBS. Then, cells were seeded into new culture bottles or plates at a certain density, and the cells that were at logarithmic phase were obtained for usage in further experimental analysis.