Cell Migration Assay with TGF-β1 and Inhibitors

A549 cells were incubated in a 24-well plate for 24 h. After serum starvation for 24 h, cells were treated with TGF-β1 and sunitinib/cRGDfK or combination for 48 h. Collected cells in culture medium containing 5% FBS was added to the bottom of a Boyden chamber. After placing over the gelatin-coated membrane filter, the silicone gasket, and the top chamber, the cell suspension was added to the top chamber, followed by incubation at 37°C in 5% CO₂ for 6 h. The membrane filter was stained using the Diff-Quick staining kit (Dade Behring) and filter was dried and stabilized on a glass slide using 30% glycerol solution, the migrated cells were counted in three randomly selected fields at 400× magnification. This experimental procedure is referred to our previous study [18 (link)].