PARPtrap™ Assay Kit for PARP1

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Catalog #
80584
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Description

The PARPtrap™ Assay Kit for PARP1 is designed to measure PARP1/DNA complex formation in a high throughput screening assay using fluorescence polarization (FP). PARP1 is known to bind damaged DNA through its DNA-binding domains. Binding to DNA activates PARP1 and in the presence of NAD+ PARP1 ribosylates itself (auto-ribosylation), what in consequence leads to PARP1 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 for PARP1 is the fluorescent-labeled oligonucleotide duplex. In the absence of ribosylation, PARP1 binds to the fluorescent probe, forming a large complex and resulting in the emission of highly polarized light. However, after auto-ribosylation, PARP1 dissociates from the oligonucleotide duplex, which then rotates freely, emitting less polarized light (Fig. 1). Addition of a PARP1 inhibitor results in trapping of PARP1 to the fluorescent oligonucleotide duplex, and increases the FP signal in a dose dependent manner.

The PARPtrap™ Assay Kit for PARP1 is a fluorescence polarization homogeneous assay. The FP signal is measured using a fluorescent microplate reader capable of measuring fluorescence polarization.

PARP1

Figure 1. PARPtrap™ Assay Kit for PARP1 schematic

Need us to run inhibitor screens or profile your compounds against PARPtrap™ for PARP1? Check out our PARP/PARPTrap™ Screening Services or DNA Replication and Repair Screening Services.

This product has been cited 4 times.

Synonyms
poly (ADP-ribose) polymerase 1, poly(ADP-ribosyl)transferase, ADPRT, PARP1, 80584-1, 80584-2, PARP-1
Product Info
Storage and Usage
Citations4
Assay Kit Format
Fluorescence Polarization
Supplied As
The PARPtrap™ Assay Kit for PARP1 comes in a convenient 96-well or 384-well format, with purified PARP1 enzyme, fluorescent-labeled oligonucleotide duplex, and PARPtrap™ assay buffer for 100 or 400 enzyme reactions.
Materials Required But Not Supplied

- Fluorescent microplate reader capable of measuring fluorescence polarization
- Adjustable micropipettor and sterile tips
- Rotating or rocker platform

Format

96 Reactions

Catalog # Name Amount Storage
80501 PARP1, GST-tag* 1 µg -80°C
78273 Fluorescent labeled oligonucleotide duplex (25 nM) 100 µl -80°C
  5x PARPtrap™ assay buffer 2 x 1 ml -80°C
  10x NAD+ 500 µl -80°C
79685 Black 96-well plate 1 Room Temp

384 Reactions

Catalog # Name Amount Storage
80501 PARP1, GST-tag* 2 x 1 µg -80°C
78273 Fluorescent labeled oligonucleotide duplex (25 nM) 2 x 100 µl -80°C
  5x PARPtrap™ assay buffer 4 x 1 ml -80°C
  10x NAD+ 2 x 500 µl -80°C
79685 Black 96-well plate 1 Room Temp

*The initial concentration of enzyme is lot-specific and will be indicated on the tube containing the protein.

UniProt #
P09874
References

1. Murai, J., et al. Molecular Cancer Therapeutics 2014. 13:433-443.
2. Murai, J., et. al. Cancer Research 2012. 72:5588-5599.
3. Zandarashvili, L., et al. Science 2020. 368(6486): eaax6367.