2.11. Immunofluorescence staining for P53

Immunofluorescence was used to analyze the expression patterns of P53 in cells. Briefly, treated cells were permeabilized by incubation with 100% methanol at 4°C for 10 min. Next, cells were rinsed with PBS, blocked with goat serum for 2 h, incubated with P53 primary antibody overnight at 4°C, and then incubated with Alexa488-conjugated secondary antibody (Invitrogen) for 1 h. Nuclei were counterstained with DAPI for 5 min, and fluorescence images were acquired using a confocal microscope. Minimum three images were used for the quantification of fluorescence intensity by Metamorph Offline software (Molecular Devices).