4T1 cells (1 × 105) were seeded in 12‐well plates for a cover density of 70–80% reached, followed by scratch making using a 200‐μl pipette. After that, cells were cultured in a serum‐free medium and treated with MATT formulations at a MATT concentration of 5 μg/ml. MATT‐LTSLs were preheated at 42°C in a water bath for 1 hr. Images were taken at 0 and 20 hr postscratching with an optical microscope (Olympus IX53, Japan).
Transwell assays were performed in 24‐well Transwell chambers with an 8‐μm pore (Corning Life Sciences, Inc.), as described in our reports.15 , 55 In brief, 4T1 cells seeded at a density of 1 × 105 cells per well in the upper chamber for 24 hr, were incubated with 200 μl of serum‐free medium and treated with MATT formulations at a MATT concentration of 1 μg/ml for 4 hr at 37°C. MATT‐LTSLs were preheated at 42°C in a water‐bath for 1 hr before incubation with cells. Subsequently, the cells in the upper chamber were removed with a cotton swab and, meanwhile, the cells attached on the lower surface of the filter were fixed with 4% paraformaldehyde, stained with 0.1% crystal violet for 10 min and washed. The stained cells were imaged by optical microscopy (Olympus IX53, Japan). Crystal violet was dissolved in 33% acetic acid and the optical density ratio was measured at the wavelength of 570 nm using a microplate reader (Multiskan FC, Thermo Fisher Scientific).
Transwell assays were performed in 24‐well Transwell chambers with an 8‐μm pore (Corning Life Sciences, Inc.), as described in our reports.