2.10. Cell migration and invasion assay

Cell migration and invasion assays were performed using modified 2‐chamber plates with a pore size of 8 μm. For migration assay, 5 × 10⁴ cells suspended in 200 μL of serum‐free medium were seeded on the upper compartment of 24‐well transwell culture chamber, and 600 μL of complete medium was added to the lower compartment. After 12‐hour incubation at 37°C, cells were fixed with methanol. For invasion assay, 30 μL of 1:8 diluted Matrigel (BD Biosciences) in serum‐free medium was added to the upper compartment of 24‐well transwell culture chamber. On the upper compartment, 6 × 10⁴ cells suspended in 200 μL of serum‐free medium were seeded and 600 μL of complete medium was added to the lower compartment. After 24‐hour incubation at 37°C, cells were fixed with methanol. Migrated or invaded cells on the lower side of the filter were stained with crystal violet and counted.