Cells have evolved DNA repair mechanisms to maintain their genetic information unaltered and a DNA damage response pathway that coordinates DNA repair with several cellular events. Despite a clear role for DNA damage in the form of DNA double-strand breaks (DSBs) in several cellular processes, the most commonly used methods to detect DNA lesions are indirect, and rely on antibody-based recognition of DNA damage-associated factors, leaving several important questions unanswered. Differently, here we describe DNA damage In situ ligation followed by Proximity Ligation Assay (DI-PLA), that allows sensitive detection of physical DSBs in fixed cells, through direct labeling of the DSBs with biotinylated oligonucleotides, and subsequent signal amplification by PLA between biotin and a partner protein in the proximity of the DNA break.
Keywords: DI-PLA; DNA damage; DNA damage response (DDR); DNA double-strand break (DSB); Imaging; PLA; Single-cell.