Transwell chambers with 8 μm membranes

We assayed the migration ability of cells using 6.5 mm diameter transwell chambers with 8 μm membranes (Corning, USA). Twenty-four hours post-transfection, A549 and H460 cells were seeded in the upper chambers, and the bottom wells were coated with 1 mg/ml matrigel for migration assays. Media containing 10% FBS were added to the bottom chambers. After 24 h at 37°C in a 5% CO₂ humidified atmosphere, cells in the upper chamber were carefully scraped off using a cotton swab, and cells that had migrated to the basal side of the membrane were fixed in methanol, stained with hematoxylin, and counted. Each test was performed in triplicate.

We assayed the invasion ability of cells using 6.5 mm diameter Transwell chambers with 8-μm membranes (Corning, U.S.A.). Cells (5 × 10⁴) were incubated into upper chamber, and the bottom wells were coated with 1 mg/ml Matrigel for the invasion assays, whereas the RPMI-1640 medium consisting of 10% FBS was supplemented to the lower chamber. After incubation for 24 h, the cells remaining on the upper membrane were discarded with cotton swab. The membrane were fixed in 4% polyformaldehyde and stained with 0.1% Crystal Violet for 15 min at 4°C. Finally, invaded cells number were counted using photographic images.