Zebra tail amplification: Accelerated detection of apoptotic blunt-ended DNA breaks by in situ ligation

Abstract

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.