Several types of DNA cuts are used as markers of apoptosis for detection of apoptotic cellsin situ. We recently introduced a ligase-basedin situassay that is specific for a single type of DNA damage—a double-strand break of DNase I-type, bearing 5′PO4 . Here we describe a vaccinia topoisomerase I-based approach to label another type of DNA damagein situ—a double-strand break of DNase II-type, bearing 5′OH. The assay uses a new type of probe, a molecular oscillator. The probe self-assembles in solution out of a dual-hairpin oligonucleotide and vaccinia topoisomerase I. The enzyme continuously separates and religates two fluorescently labeled hairpins, which can participate in energy transfer. We describe the successful combination of topoisomerase- and ligase-based systems into anin situassay. The assay uses an oscillating probe for simultaneous detection of two types of DNA cuts in tissue sections.
