Migration and invasion assays

Cells (20,000 cells/0.5 ml/well) were plated onto control membrane inserts with 8-micron pores or Matrigel-coated membrane transwell inserts (BD Biosciences, Bedford, MA, USA), which are placed in 24-well chambers filled with 0.6 ml growth medium. Twenty-four hours after plating, cells that remained on the upper surface of the membrane were removed by cotton-tipped swabs, and cells that migrated/invaded to the lower surface of the membrane were viewed under the fluorescent microscope. The percent migration was expressed as percent migration = (mean number of cells migrating through Matrigel insert membrane × 100/mean seeding control) and percent invasion was expressed as: percent invasion = (mean number of cells invading through Matrigel insert membrane × 100/mean number of cells migrating through control insert membrane.