Boyden chambers 24 well

Boyden assays were performed using 8 µm pores Boyden Chambers (24-well, Costar). For the invasion assays, the upper chamber was coated with 6 µL of Matrigel (BD Biosciences) dissolved in 100 µL of DMEM. HeLa cells were detached and washed with DMEM 0.1% BSA. In total, 100,000 cells were seeded in the top chamber and allowed to migrate for 6 h (migration) or 24 h (invasion) toward the bottom chamber containing 10% FBS. Upper and lower chambers were then washed with 1x PBS and cells on the bottom side of the chamber were fixed with 4% PFA. Cells in the upper chambers were removed using cotton swabs and the membrane was mounted on a glass slide using SlowFade Gold reagent (Invitrogen). The average number of migrating cells in 10 independent 20× microscope fields were evaluated, and each experiment was performed in triplicate.

Boyden assays were performed using 8 µm pores Boyden Chambers (24-well, Costar). For the invasion assays, upper chamber was coated with 6 µL of Matrigel (BD Biosciences) dissolved in 100 µL of DMEM. Cells were detached and washed with DMEM 0.1% BSA. One hundred thousand cells were seeded in the top chamber and allowed to migrate for 6 h (migration) or 16 h (invasion) toward the bottom chamber containing 10% FBS. Upper and lower chambers were then washed with 1×PBS and cells on bottom side of the chamber were fixed with 4% PFA. Cells in the upper chambers were removed using cotton swabs, and the membrane was mounted on a glass slide using SlowFade Gold reagent (Invitrogen). The average number of migrating cells in 10 independents ×20 microscopic fields were evaluated, and each experiment was performed in triplicates.