Zebra Tail Amplification: Accelerated Detection of Apoptotic Blunt-Ended DNA Breaks by In Situ Ligation

In situ ligation (ISL) is a simple and specific technique for apoptosis labeling in tissue sections. In its most economical version ISL uses ordinary PCR-labeled DNA fragments as probes. In tissue sections these makeshift probes are ligated to apoptotic DNA breaks by T4 DNA ligase. The approach can selectively label 5'PO₄ DNA breaks with blunt ends, and is the histological equivalent of electrophoretic apoptotic ladder detection. The main drawback of this technique is its low speed, as it requires 18 h-incubation for efficient labeling. Here, we describe an easy modification of ISL which reduces the incubation time to 1 h and converts ISL into a rapid detection method taking ~3 h overall. Signal enhancement is achieved by a new type of isothermal amplification reaction which generates "zebra tails"- long and labeled extensions of the probes attached to DNA breaks.