Transwell polycarbonate membrane cell culture inserts

Both cell migration and invasion experiments were conducted with Transwell polycarbonate membrane cell culture inserts (BD Biosciences, CA, USA). The cells were pre-starved in serum-free medium, then single-cell suspension was prepared via trypsin digestion. The cell concentration was calculated and adjusted to 1 × 10⁵/100 μL. Each 100 μL cell solution was poured onto the upper compartment of polycarbonate Transwell chamber (Matrigel was applied for the filter for invasion assay (BD Biosciences, CA, USA)). 750 μL of complete culture medium containing 10% FBS was filled in the lower chamber as attractant. After 20 h, the free cells inside insert were completely wiped off with cotton Q-tips. The migrated/invaded cells were fixed with 4% paraformaldehyde and followed by crystal violet staining for 15 min. The cells were counted in three independent fields under optical microscope to calculate the transwell capacity.

Both cell migration and invasion experiments were conducted with Transwell polycarbonate membrane cell culture inserts (BD Biosciences, CA, USA). A total of 3 × 10⁴ cells were added to the upper chamber, and the cells were allowed to migrate to the lower chamber for 24 h. (Matrigel was applied for the filter for invasion assay (BD Biosciences, CA, USA). The migrated/invaded cells were fixed with 4% paraformaldehyde and stained with 0.1% crystal violet staining for 30 min. The cells were counted in three independent fields under an optical microscope to calculate the transwell capacity.

Cell migration and invasion assays were performed according to a previous study [18 (link)]. Corning Transwell polycarbonate membrane cell culture inserts with 8.0-μm pores were uncoated for migration assay, or coated with 50 μL of BD Matrigel Basement Membrane Matrix (BD Biosciences, USA) diluted 1:3 with FBS-free DMEM (fetal bovine serum free Dulbecco’s Modified Eagle Medium) for invasion assay. Cells were trypsinized and then resuspended in serum-free medium. 2 × 10⁴ cells were seeded in the top chamber with serum-free medium and allowed to migrate toward serum-containing medium in the lower chamber for 24 h. Then cells were fixed in 4% paraformaldehyde and stained with 0.01% crystal violet (AS1086, ASPEN, China) for 10 min. After wiping the cells remaining in the upper chamber with cotton swabs, the migratory or invasive cells were imaged and counted using a Leica AM6000 microscope (Leica Microsystems, Germany). The numbers of migratory or invasive cells were calculated as the average numbers of five random fields per well under 100× magnification.