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Boyden chamber transwell assays
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In brief, we used 24-well Boyden chambers (Corning, USA) for migration assays as previously described25. Transfected cells (KYSE30,1 × 104 or KYSE450, 5 × 104) were plated into the upper chamber and cultured in serum-free RPMI 1640 medium with or without treatment (cells overexpressing PLAU with U0126 added in the upper chamber). WT KYSE-30 and KYSE-450 cells were treated with CM of CAFs with or without recombinant PLAU in the presence or absence IPR-803. We counted the numbers of cells that migrated in five different areas at 100-fold magnification.
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