2.8. Transwell assay

The Transwell chambers were bought from CORNING. After transfection, cells were diluted into the concentration of 10⁵/well and translocated to the chambers, which were added with DMEM (free FBS). Under the chambers, DMEM containing 20% FBS was filled in the wells. After culturing for 36 hours, the medium was discarded and cells were stained by crystal violet, presenting with blue. Then, the stained cells were captured under the microscope (EVOS M7000; Thermo Fisher) at 100×. Focusing on one field, the blue spots represented the cells that traversed through the porous membrane and glue to reflect the relative invasion rate. The numbers of blue spots were counted and calculated by ImageJ (NIH).