Quantification of beta-galactosidase activity as a marker of radiation-driven cellular senescence

Cellular senescence is a permanent state of cell cycle arrest that can be triggered by different stressors, including cancer treatments (the so-called “therapy-induced senescence”), such as radiation therapy (RT). Although senescent cells do not proliferate, they remain metabolically active and play a critical role in tumor progression, metastasis, and response to therapy. Therefore, investigating the induction of cellular senescence upon RT treatment is a critical read out for investigating RT efficacy or combinatorial strategies in cancer research. Senescent cells are characterized by a plethora of markers, including an increased content and activity of lysosomes, which can be detected by the activity of the lysosomal enzyme senescence-associated β-galactosidase. In this chapter, we present a protocol for the gold standard cytochemical method for quantification of the activity of the senescence-associated β-galactosidase in irradiated murine breast cancer cells in vitro.