Transwell chambers with 8 μm pore membranes

The experiment was performed using transwell chambers with 8 μm pore membranes (Millipore). For the migration assay, cells were seeded into the top chamber in FBS-free medium at a density of 8×10⁴ cells per chamber, and 600 μL of 10% FBS-containing medium was placed in the bottom chamber as a chemoattractant. For the invasion assay, 8×10⁴ cells per chamber were plated on the top chamber with a Matrigel-coated membrane. After 48 h, the cells were stained with 0.1% crystal violet solution and counted under an inverted microscope (Leica).

The experiment was performed using Transwell chambers with 8 μm pore membranes (Millipore). For the migration assay, cells were seeded into the top chamber in FBS-free medium at a density of 5 × 10⁴ cells per chamber, and 600 μl of 10 % FBS-containing medium was placed in the bottom chamber as a chemoattractant. For the invasion assay, 10⁵ cells per chamber were plated on the top chamber with Matrigel-coated membrane. After 24 h or 48 h, the cells were stained with 0.05 % crystal violet solution and counted by an inverted microscope (Leica).

Cell invasion was detected using the Transwell invasion assay. Transwell chambers with 8-μm-pore membranes (Millipore, Bedford, MA, USA) were coated with Matrigel (BD Biosciences, San Diego, CA, USA) according to the manufacturer’s instructions. A total of 200 μl of serum-free medium containing 1 × 10⁵ cells was added to the upper chamber, and 600 μl of medium containing 10% FBS was added to the lower chamber. The cells were cultured for 24 h at 37°C, noninvasive cells on the top surface were removed by a cotton swab, and the invasive cells on the lower surface were fixed with 4% formaldehyde and stained with 0.1% crystal violet. Then cells were observed and counted under the microscope.