Fiji-Based Tool for Rapid and Unbiased Analysis of SA-β-Gal Activity in Cultured Cells

Normal cells under stressful conditions such as DNA damage or excessive mitogenic signaling may undergo senescence, which is associated with cell cycle arrest and induction of a proinflammatory phenotype. Accumulation of senescent cells may contribute to the shortening of the life span by accelerating aging and promoting chronic diseases. Cytochemical detection of the senescence-associated β-galactosidase (SA-β-gal) activity with 5-bromo-4-chloro-3-indolyl β-D-galactopyranoside (X-gal) is a widely recognised marker of cell senescence. However, its simplicity and cost effectiveness lead to limitations in quantification, which is usually limited to manual counting of the positive cells. In order to address those limitations, we developed a Fiji-based macro extension that performs automatic and unbiased analysis of the integrated density of SA-β-gal specific signal. Our tool is not only faster than manual counting but also provides extra resolution compared to the manual methods. Our macro extension could be a valuable tool in any senescence research laboratory.