4.6. Senescence-Associated β-Galactosidase Activity

SA-b-gal was assayed 10 days after irradiation (0, 2, 4 and 10 Gy) according to previously describe protocol [22]. Cells were fixed with 5% formalin for 5 min, wash with PBS and stained with fresh β-galactosidase staining solution (1 mg/mL 5-bromo-4-chloro-3-inolyl-β-galactosidase in dimethylformamide (20 mg/mL stock), 5 mM potassium ferricyanide, 150 mM NaCl, 40 mM citric acid/sodium phosphate and 2 mM MgCl₂, at pH 6.0). Cells were incubated overnight at 37 °C and bright-field pictures of the cells were taken with EVOS microscope (Life Technologies, Burlington, ON, CA).