The scratch assay was applied to evaluate cell migration and repair. After reaching 90%–100% confluency in wells of culture plates, cells were exposed to serum-free medium for 6 h, and each cultured well was scraped with a pipette tip in the same specification. Cells were washed with PBS to remove fragments. Microscope images of the same positions were acquired in after 0 and 30 h. Based on the percentage of wound closure area, cell migration was determined.
Transwell migration and invasion assays were conducted to evaluate the ability of cell migration and invasion. 24-well transwell chambers (Corning, United States) were used in the assay. For the invasion assay, matrigel (Corning, United States) was applied to the upper ventricle surface of the basement membrane of the transwell chamber. The insert was filled with 30,000 cells suspended in 150 ul serum-free serum before the assay. In the lower chamber, 700 ul medium containing 12% fetal serum was added for chemotactic stimulation. Cells were cultured for 24 h for migration assays and 40 h for invasion assays. Then cotton swabs were used to remove cells from the surface of the membrane. Cultured cells were fixed with 100% methanol and stained with 0.1% crystal violet. Random visual fields of 3 different inserts were captured, and the number of cells was counted.
After inoculating 3000 cells per well, Huh7 and Hep3B cells were grown for 8 and 10 days respectively in 6 well plates in complete medium. Cultured cells were fixed with 100% methanol and stained with 0.1% crystal violet. Each well was counted for the number of colonies.
Transwell migration and invasion assays were conducted to evaluate the ability of cell migration and invasion. 24-well transwell chambers (Corning, United States) were used in the assay. For the invasion assay, matrigel (Corning, United States) was applied to the upper ventricle surface of the basement membrane of the transwell chamber. The insert was filled with 30,000 cells suspended in 150 ul serum-free serum before the assay. In the lower chamber, 700 ul medium containing 12% fetal serum was added for chemotactic stimulation. Cells were cultured for 24 h for migration assays and 40 h for invasion assays. Then cotton swabs were used to remove cells from the surface of the membrane. Cultured cells were fixed with 100% methanol and stained with 0.1% crystal violet. Random visual fields of 3 different inserts were captured, and the number of cells was counted.
After inoculating 3000 cells per well, Huh7 and Hep3B cells were grown for 8 and 10 days respectively in 6 well plates in complete medium. Cultured cells were fixed with 100% methanol and stained with 0.1% crystal violet. Each well was counted for the number of colonies.
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