Serum-free suspension (2×105 cells /ml) was seeded in a Transwell chamber either pre-coated with Matrigel or not. Serum-contained medium was added to the bottom as an inducer. After 24-h cell culture, cells penerated to the bottom were fixed, and dyed in 0.2% crystal violet. They were captured in 5 random fields per well for cell counting.

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