Effect of Aurora kinase inhibitors on the mitotic phosphorylation of Borealin. The mobility shift of Borealin was determined by Western blotting using a 12.6% SDS-polyacrylamide gel. Transfers were stripped and reprobed for β-actin to serve as loading control. (A) The response of cells to different doses of ZM447439. Hela cells were treated for 16 hours with the indicated doses of ZM447439 to block Aurora B kinase activity and then stained to detect DNA using immunofluorescence. The percentage of normal, abnormal and multinucleated cells was determined using fluorescence microscopy. (B) Effect of ZM447439 pre-treatment on mitotic phosphorylation of Borealin. WT-8 cells were treated with ZM447439 for 1 hour and then blocked in mitosis by treating for 16 hours with nocodazole (ZM447439 was left on during the nocodazole treatment). DMSO was used as a vehicle control. (C) Effect of high dose of ZM447439 and VE465 on mitotic phosphorylation of Borealin. WT-8 cells treated with 10 μM ZM447439 or 100 nM VE465 for 1 hour were either left to grow asynchronously or blocked in mitosis by treating for 16 hours with nocodazole (aurora kinase inhibitors were left on for the duration of the experiment). DMSO was included as a vehicle control.