Transwell pore polycarbonate membrane insert

Transwell pore polycarbonate membrane inserts (Corning, pore size 8 μm, MD, USA) were used, and a layer of Matrigel (Corning) was coated on the insert during the invasion experiment. Cells were inoculated inside the chamber with or without exosome treatment. The concentration of exosomes was 50 μg/mL 18 (link), 19 (link). The upper chamber used RPMI-1640 medium, and the lower chamber used RPMI-1640 medium containing 20% FBS medium with or without exosomes. After 48 h of culturing, the cells were fixed and stained. Finally, the residual crystal violet was washed away with water. Cells that passed through the chamber were counted.

Migration assay was performed using Transwell pore polycarbonate membrane insert (Corning, New York, USA), invasion assay was performed using Matrigel-coated invasion chambers (BD Biocoat, Corning). Cells were seeded into the upper chamber coated with matrigel, with or without exosome treatment, and cultured in serum-free medium. The lower chamber contained medium supplemented with 20% normal or exosome-free fetal bovine serum. Cells invading the membrane and adhering to the lower surface after 24 h were fixed with methanol and then stained with crystal violet for 20 min. Cells adhering to the lower surface were then washed with PBS and cells on the upper membrane were removed. Images of the stained cells were captured using a microscope (Nikon).