Matrigel invasion chambers with 8 μm pore inserts

The cell invasion assay was performed using six-well Matrigel invasion chambers with 8-μm pore inserts (BD Biosciences). Cells (5 × 10⁴) cells were seeded into the upper chambers. The lower chambers were filled with serum-free medium. After 24 h, the cells in the upper chamber were treated with HuR-FNP in serum-free medium. After 6 h of transfection, the culture medium in the upper chamber was replaced with 2% serum-containing medium with or without AMD3100 (100ng/ml), while the lower chamber was replaced with 20% FBS-containing medium and incubation continued. After an additional 48 h of incubation, the filters were removed, processed and the number of invaded cells counted as previously described [34 (link)–36 (link)]