Cell migration and invasion assay

Wound healing assay and transwell assay were performed to measure the migration and invasion ability of breast cancer cells respectively. For wound healing assay, when A549 CSC and H1299 CSC cells were cultured to 90% confluence in 96-well plates, the medium was removed and a gap was made by enforcing the sterile pipette tip on the monolayer cells. The width of the wound gap at 24 h was acquired and normalized to initial distance at 0 h. Migration rate was calculated using the following formula: migration rate = migration distance/ original distance. For transwell assay, A549 CSC and H1299 CSC cells were suspended in 200 ml serum-free DMEM and seeded in chambers (8 mm, BD Biosciences) coated with BD BioCoat Matrigel. After incubation, the non-invaded cells on the upper membrane surface were removed with a cotton tip. The cells on membrane were fixed and stained by violet crystalline.