Cell Migration and Invasion Assays

Cell migration and invasion assays were performed as previously described (20 (link)). Transwell chambers (Corning, Corning, NY, USA) equipped with 8-μm pore insets were used for the migration and invasion assays. For the migration assay, Transwell chambers (Corning, Corning, NY, USA) containing 8-μm pores were uncoated with Matrigel in the upper chamber. For the invasion assay, Transwell chambers (Corning, Corning, NY, USA) containing 8-μm pores were coated with 100 μl of 1:8-diluted Matrigel (BD Biosciences) in the upper chamber. Briefly, 100 μl of cell suspension (8 × 10⁴ cells) of serum-free medium was plated in the upper chamber; medium with 10% FBS was added to the lower chamber. After incubating 18 h for migration and 22 h for invasion, we removed the cells of the upper chamber and then stained the migration and invasive cells of the lower chamber using crystal violet solution. The results were photographed by light microscopy and counted by ImageJ. The experiment was done in triplicate.