Cell Migration and Invasion Assays

BD Falcon cell chambers (BD biosciences, CA, USA) were employed to evaluate cell migration. 2.5 X 10⁴ cells in 500 μL serum free DMEM were added to each transwell insert without matrigel. 500 μL NIH-3T3 conditioned media was added to the lower chambers and cells were cultured for 20 h at 37°. BD Falcon Matrigel invasion chambers (BD biosciences, CA, USA) were used to measure cell invasion following the manufacturer’s instruction and with modifications as described [18 (link)]. For both assays, samples were done in quadruplet for 2 experiments. Membranes without and with Matrigel were removed from inserts, and then washed, fixed with 100% methanol, and stained with 1% Toluidine blue. Each membrane containing cells were counted using a Nikon TMS microscope. An Olympus IX73 was used to image the cells, to determine the number of either migratory or invasive cells per well.