The PARPtrap™ Assay Kit for PARP2 is designed to measure PARP2/DNA complex formation in a high throughput screening assay using fluorescence polarization (FP). Similarly, to PARP1, PARP2 recognizes and binds damaged DNA via its DNA-binding domain. Binding to DNA activates PARP2 and in the presence of NAD+ PARP2 ribosylates itself (auto-ribosylation), which leads to PARP2 dissociation from the DNA due to the accumulated negative charge of the ribosyl polymer. In the presence of some inhibitors, however, PARP remains bound to the DNA, a phenomenon termed trapping. Trapped PARP-DNA complexes have been shown to be highly cytotoxic to cancer cells, therefore such inhibitors may be desirable for cancer treatment.

The key to the PARPtrap™ Assay Kit is the fluorescent-labeled oligonucleotide duplex. In the absence of ribosylation, PARP2 binds to the fluorescent probe, forming a large complex and resulting in the emission of highly polarized light. However, after auto-ribosylation PARP2 dissociates from the oligonucleotide duplex, which then rotates freely, emitting less polarized light (Fig. 1). Addition of a PARP2 inhibitor results in trapping of PARP2 to the fluorescent oligonucleotide duplex, and increases the FP signal in a dose dependent manner.

The PARPtrap™ Assay Kit is a fluorescence polarization homogeneous assay. The FP signal is measured using a fluorescent microplate reader capable of measuring fluorescence polarization.

Figure 1. PARPtrap™ Assay Kit for PARP2 schematic