Wound healing, transwell migration and invasion assays

For wound healing assay, the cells were grown to a confluent monolayer, following which a line was scratched through the monolayer in each well and non-serum media was added. Three representative wound lines were selected for measurement. The wound areas were visualized and measured at 0 and 24 h post-wounding.

In the migration assay, 5 × 10⁴ cells were seeded into the upper chamber of the transwell plate (Millipore, La Jolla, CA, USA). The cell invasion assay was performed using Matrigel-coated filters (BD, La Jolla, CA, USA). Specific culture medium was added to the bottom chamber. As previously described, BC cells were allowed to migrate for 24 h or invade through the Matrigel for 48 h [60]. The migrated or invaded cells were then fixed and stained with 0.1% crystal violet, six randomly selected fields were photographed, and the cell numbers were counted.