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Interrogating TREX1 with the Transcreener dAMP Exonuclease Assay

Enzolution TREX1 Assay System

The Enzolution TREX1 Assay System includes TREX1 enzyme, DNA substrate, assay plates, and buffer components required for a TREX1 reaction to produce dAMP. The Transcreener dAMP Exonuclease Assay (sold separately) measures dAMP to determine enzymatic activity. These measurements allow researchers to screen compound libraries for modulators to help find new disease therapies.

The Transcreener dAMP Exonuclease Assay is not included in the Enzolution Assay System but is available for separate purchase through BellBrook Labs. For dAMP assay ordering information, please view Transcreener dAMP Exonuclease Assay.

The Comprehensive Approach to Measure TREX1 Enzymatic Activity

Enzolution TREX1 Assay System Schematic

Enzolution Assay System includes enzyme, substrate, & buffers for a TREX1 reaction. Transcreener dAMP Assay measures the dAMP produced.

Robust Assay Yields Quality Data

The Enzolution TREX1 Assay System used with the Transcreener dAMP assay produces robust results amenable to HTS. Z’ measurement using optimized TREX1 assay conditions (n=16). Z’ = 0.88.

Screen for Inhibitors & Perform SAR

The Enzolution TREX1 Assay System used with the Transcreener dAMP assay is designed to screen compound libraries in a high throughput format to identify potential inhibitors. Follow-up SAR can also be performed using the assay to determine inhibitor potency with ease.

Trex1 dose-response curves v3

The Transcreener dAMP Exonuclease Assay allows selectivity profiling for TREX1. Suramin IC50 = 3.0 µM.

Detection of dAMP to Measure TREX1 Activity - FP Readout

The enzyme buffer includes 10 mM Tris pH 7.5, 7.5 mM MgCl2, 0.005% BSA, and 0.01% Brij-35. Here we use 35 nM ISDna. The Detection mix included 20 μg/mL dAMP antibody, 2 nM dAMP Alexa 633 tracer, and 1X Stop & Detect Buffer B in 10 μL. The TREX1 enzyme reaction took place for 60 minutes at 30°C. The Detection Mix was added and incubated for 90 minutes at room temperature and read with a CLARIOstar plate reader. Linearity is shown here when raw data is converted to dAMP formed using a standard curve.

TREX1 Enzyme Titration

TREX1 Assay Enzyme Titration

Linear Response

TREX1 Assay Enzyme Titration Product Formed

Overview of the Transcreener dAMP Exonuclease Assay (Sold Separately)

Mix-and-Read-GEF-Assay

The Transcreener dAMP Exonuclease Assay directly measures dAMP produced during the biological functions like the degradation of DNA. Transcreener dAMP technology uses polyclonal mouse antibodies that selectively bind to dAMP and a far-red fluorescent tracer. dAMP produced displaces the tracer, changing the fluorescent properties and providing a readout.

It is an easy-to-use, mix-&-read, HTS-ready assay that yields robust results. All you need to do is perform the enzyme reaction, add Transcreener reagents, and read your plates. It is available with an FP fluorescent readout.

BellBrook's Recombinant TREX1 Enzyme

Active, human Three Prime Repair Exonculease 1 enzyme, His-tagged protein expressed and purified from baculovirus-infected insect cells. The enzyme has been thoroughly validated with Transcreener dAMP Exonculease Assay Kit. For specific activity, please refer to the Certificate of Analysis for each individual enzyme lot. Purity is >90% in a 50 mM Tris, 500 mM NaCl, and 20% glycerol buffer composition (pH 8.0). Currently used with the Transcreener assay at a concentration between 0.06 - 0.52 nM in a 10 μL reaction.

TREX1 Enzyme Schematic v2

TREX1 Assay Services

Interested in moving your program forward, but don't want to bring an assay in-house? Our scientists can help! BellBrook scientists will use their extensive biochemistry and enzymology expertise to work with you and accelerate your TREX1 program.

Assay Development Services

Lead Discovery Services Include:

  • Inhibitor Screening – To identify or confirm activity with the target.
  • Inhibitor Potency Profiling – Dose-response with target and/or related proteins. Fast IC50 results.
  • Residence Time Measurements – Determination of koff using ‘jump dilution’ enzymatic assay method.
  • Mechanism of Action Studies – Kinetic analysis to define the mode of inhibition.

Contact Us to Learn More

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What's Included

What You Will Need

Enzolution TREX1 Assay System

* For custom or bulk orders (over 100,000 wells) please contact us (info@bellbrooklabs.com) for a quote.

Transcreener dAMP Exonuclease Assay Kits

* For custom or bulk orders (over 100,000 wells) please contact us (info@bellbrooklabs.com) for a quote.

TREX1 Enzyme Ordering Information

* For custom or bulk orders (over 100 µg) please contact us (info@bellbrooklabs.com) for a quote.

Related Resources

Interconnections Between the DNA Damage Response and Innate Immunity

There is extensive crosstalk between the DNA damage response (DDR) pathways and innate immune pathways. Both the individual pathways and the interconnections between them are a focus for exciting new small molecule drug therapeutics that target cancers and debilitating autoimmune disorders. The Transcreener HTS Assay platform accelerates these efforts by providing a robust and easy-to-use biochemical assay to measure activity of key enzymes in the innate immune and DDR pathway.

In this guide, we provide an overview of the DDR and innate immune pathway, and describe Transcreener Assays and Assay Systems for key therapeutic targets. The enzyme targets discussed include:

DDR

  • POLQ
  • WRN
  • PARP1
  • PARG
  • DNA PK

Innate Immunity

  • cGAS
  • TREX1
  • TBK1
  • IKKβ
  • IRAK4
  • ENPP1
  • CD38
  • OAS-1
  • RIG-1/MDA5
  • DDX3
  • CD39
  • CD73
DNA Damage Response and Innate Immunity Preview

TREX1 As a Therapeutic Target

TREX1 is a crucial component of the innate immune response acting as the main exonuclease responsible for degrading cytosolic DNA. Reduction of cytosolic DNA leads to inactivation of the cGAS/STING Pathway, which is essential for intrinsic antitumor immunity. As a result, TREX1 plays a pro-tumorigenic role in cancer, and its inactivation is a promising strategy for future monotherapy and combination treatments.

The Discovery of TREX1 antagonists to reign in overactivity requires selective technologies that can help assess the ability to interact with the target. Robust, HTS-ready assays that are accurate yet flexible provide a powerful tool to accelerate discovery. 

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