Transwell biocoat growth factor reduced matrigel invasion chambers

Twelve-well cell culture insert, and 24-well Transwell BioCoat growth factor-reduced Matrigel invasion chambers (BD Biosciences, 8 μM pore size) were used for migration and invasion assays, respectively. After overnight serum starvation, cells were plated to the upper side of the Transwell device, in triplicates, in serum-free medium, whereas the lower well contained regular 10% FBS culture medium to create an FBS gradient. We plated 80,000 cells for the migration assay and ended the experiment after 5 or 14 h for SKOV3 and IGROV-1, respectively (according to their respective migratory capacities). For the invasion assay, we plated 50,000 cells and stopped the experiment after 24 or 48 h for SKOV3 and IGROV-1, respectively (here again, defined according to their respective migratory capacities). At the end of the experiment, the remaining cells in the upper side of the Transwell device were removed. Migrating and invading cells at the bottom side of the Transwell device were fixed and stained with crystal violet for 30 min and then counted in five different representative fields (× 5 objective, Zeiss Axioplan microscope, AxioCamERc 5s).

Twenty-four-well Transwell BioCoat™ growth factor-reduced Matrigel™ invasion chambers (BD Biosciences, 8 μM pore size) were used for the invasion assay. After overnight serum starvation, cells were plated to the upper side of the Transwell device in serum-free medium, whereas the lower well contained regular 5 % horse serum (MCF10A) or 10 % FCS (MDA-MB-231, CAMA-1) culture medium to create a serum gradient. We seeded 5 × 10⁴ cells and stopped the experiment 48 h later. The remaining cells in the upper side of the Transwell device were removed and the invading cells at the bottom side of the Transwell device were fixed and counted.