LysA™ Universal PARylation Assay Kit

Catalog #

82123

$1,100 *

Size: 96 reactions

Qty

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*US Pricing only. For international pricing, please contact your local distributor.

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Description

LysA™ Universal PARylation Assay Kit is a sandwich ELISA-based kit designed to measure and quantify the amount of total poly ADP-ribosylation present in cell extracts. The kit comes in a convenient 96-well format and contains all the reagents needed, including a PAR standard that allows to establish a standard curve for quantitative measurements. It contains enough antibodies, blocking buffer, and detection reagents to measure PARylation levels in cell extracts, in a 96 well plate format. It also includes cell lysates as controls for assay performance.

Note: The assay is adequate for Poly-ADP-ribosylation detection only. The assay is linear in the 100 pM to 20 nM PARylation range.

Figure 1: LysA™ Universal PARylation Assay workflow diagram.
An anti-PAR antibody is used to coat the 96-well plate. Lysates from cells are added to the coated wells, so PAR (PARylated proteins) present in the cell lysates are captured by the antibody. This is followed by an incubation with an anti-PAR primary antibody and a secondary HRP-labeled antibody. Addition of a chemiluminescent HRP substrate provides a luminescence signal, which directly correlates with the amount of PAR present in the cell extracts

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Synonyms

PARP1, PARP2, PARG

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Product Data Gallery

Example of PAR standard curve.

Effect of the PARG inhibitor PDD00017273 on cellular PARylation levels in various cell lines.

Effect of various PARP inhibitors on cellular PARylation levels in HEK293 cells.

Product Info

Storage and Usage

Citations

Assay Kit Format

Chemiluminescent

Materials Required But Not Supplied

Modified RIPA Lysis Buffer (Moderate Strong) (BPS Bioscience #82126)
LysA™ Protease Inhibitor Cocktail Kit (BPS Bioscience #82199)
ADP-Ribosylation Cycle Inhibitor Mix (BPS Bioscience #82130)

Format

Catalog #	Name	Amount	Storage
	10 µM PAR Standard	6 μl	-80°C
	Anti-PAR Coating Antibody	10 μl	-80°C
	Anti-PAR Detection Antibody	10 μl	-80°C
82156	HEK293 High PAR Cell Lysate	300 µl	-80°C
82157	HEK293 Low PAR Cell Lysate	300 µl	-80°C
52131H	Secondary HRP-Labeled Antibody 2	10 μl	-80°C
79743	Blocking Buffer 3	2 x 25 ml	+4°C
79670	ELISA ECL Substrate A	6 ml	Room Temp
79670	ELISA ECL Substrate B	6 ml	Room Temp
79837	96-well module Maxisorp™ plate		Room Temp

Background

PAR homeostasis is regulated by the family of PAR polymerases (PARPs) and PARG (Poly (ADP-ribose) glycohydrolase) in response to cellular stress conditions. ADP ribosylation, which is the addition of an ADP-ribose to a protein, is a reversible post-translational modification of proteins mostly involved in the DNA Damage Response (DDR) pathway. Poly-ADP-ribosylation (termed PARylation) is the addition of linear or branched chains of ADP-ribose. PARP and PARG activity are linked to cellular responses in inflammation, ischemia, stroke, and cancer. PARP inhibitors have been used in cancer treatment with success, leading to synthetic lethality when homologous recombination repair (HRR) is already defective. PARG is overexpressed in breast cancer and associated with tumor growth and survival. Decrease in PARG activity can potentiate the effect of current cancer therapies, such as chemotherapy and radiation, making PARG inhibition with selective inhibitors a promising approach in cancer and immunotherapy. Further studies of PARP and PARG will elucidate how the levels of PARylated proteins contribute to disease and can be modulated to provide therapeutic benefit.

References

Marques M., et al., 2019 Oncogene 38 (12): 2177-2191.
James D. I., et al., 2016 ACS Chem Biol 11 (11): 3179-3190.
Drown B. S., et al., 2018 Cell Chem Bio 25 (12): 1562-1570.

Storage/Stability

This assay kit will perform optimally for up to 6 months from date of receipt when the materials are stored as directed.

Applications

Quantify total levels of PAR in cultured cells.
Screen or determine the IC₅₀ of PARP or PARG inhibitors in cultured cells.
Validation of PARP or PARG inhibitors in a cellular model.

Shipping Temperature

-80°C

Notes

Troubleshooting Guide
Visit bpsbioscience.com/assay-kits-faq for detailed troubleshooting instructions. For all further questions, please email [email protected]

Contraindications

This assay does not allow detection of mono-(ADP-ribose) (MAR) modifications on proteins.
Avoid SDS or other strongly denaturing detergents at concentration > 0.1%.
This assay is compatible with up to 2% v/v total detergent present and up to 1% DMSO.
Avoid the use of sodium azide and reducing agents, such as dithiothreitol or β-mercaptoethanol at concentration >10 mM.

Warnings

Avoid freeze/thaw cycles

Disclaimers and Limitations

BPS Bioscience assay kits are validated using the listed components according to our specific protocol. Any deviations to kit components or protocols, such as using different proteins, cell/tissue lysates, or buffers (including using the same component from other commercial sources) will void any warranties on the performance of the assay kit and is not recommended.

End-users should immediately report any issues to [email protected] upon receipt of the kit, such as missing components, damaged vials, no dry ice, etc.

Scientific Category

Cancer Therapy Target

Regulatory Status

Research Use Only