Treated Hela cells on coverslips were washed once with prewarmed PBS and fixed in 4% (w/v) paraformaldehyde for 15 min. After three washes in PBS, cells were permeabilized with 0.2% (v/v) Triton X-100 for 5 min. After three washes in PBS, cells were blocked in PBS containing 5% (w/v) bovine serum albumin (BSA) for 30 min and incubated with indicated antibodies in PBS containing 3% (w/v) BSA for 2 hours at 37°C. After three washes, cells were incubated with Alexa Fluor 488–conjugated secondary antibodies for 1 hour at 37°C and then with 4′,6-diamidino-2-phenylindole (Roche) for 15 min. The coverslips were washed extensively and mounted onto slides. Imaging of the cells was carried out using an Olympus BX51 microscope confocal system under a 40× objective.

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