Transcreener® ADP² AMPK Assay

Learn More About Transcreener Method Used for This AMPK Assay

Discover AMPK Modulators with the Transcreener ADP2 Assay

The Transcreener ADP² Kinase Assay directly measures the ADP produced by  AMPK (also known as AMP-Activated Protein Kinase or 5' Adenosine Monophosphate-Activated Protein Kinase). By measuring the production of ADP researchers can effectively determine the activity of the AMPK enzyme. The assay provides a powerful tool to screen compound libraries for AMPK modulators to help find new therapeutics for a variety of different diseases including coronavirus, autoimmune diseases, and cancer.

The Role of AMPK As Therapeutic Target

AMPK is a drug target in many areas, including metabolism, inflammatory diseases, tumor suppression, and more recently, the coronavirus. SARS-CoV-2 uses the ACE2 receptor to infect the human body. Research has shown that AMPK can increase the expression and stability of ACE2. Once the coronavirus is in the body, it leads to a downregulation of ACE2. This causes a chain of events leading to deadly cardio-pulmonary issues. One drug currently on the market Metformin, activates AMPK via phosphorylation. Currently used as a Diabetes drug, Metformin has positive effects on both lipid and glucose metabolism via AMPK. Another AMPK activator AICAR has shown to improve alveolar and capillary barriers preventing damage from an infection like SARS-CoV-2 in mice. Further work needs to be done but AMPK is a target of interest for coronavirus treatment.

Direct Detection of ADP to Measure AMPK Enzymatic Activity

ADP FP Kinase Assay

The workhorse. Used in many large screens. Best deck and signal stability.

ADP FI Kinase Assay

Positive FI signal. Compatible with simple fluorescence plate readers. Faster read time than FP or TR-FRET.

ADP TR-FRET ATPase Assay

For customers who prefer TR-FRET detection. Uses the same filter set as HTRF®.

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Learn why direct ADP detection is the best HTS assay method for Kinases and ATPases

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    Applications

    • Measure Enzymatic Activity of AMPK
    • Screen Compound Libraries for AMPK Modulators
    • Quantify Inhibitor Potency
    • Inhibitor Selectivity Profiling
    • Measure Drug-Target Residence Time

    Features

    • Direct detection of unlabeled ADP
    • Easy to use, homogenous, one-step format
    • Robust Assay Z’ > 0.7 under initial velocity conditions
    • Far-red fluorescent readouts minimize compound interference
    • A safe, non-radioactive method
    • Available in FP, FI, or TR-FRET readouts

    Easy-to-Use, Mix-and-Read, HTS-Ready Assay

    The Transcreener ADP² AMPK Assay is in a simple, HTS-ready, mix-and-read format. Run your enzyme reaction, add Transcreener reagents, and read your plates. The assay is compatible with 96, 384, and 1536-well formats.

    Detection of ADP Under AMPK Initial Velocity - FP Readout

    The assay demonstrates linearity when raw data is converted to ADP using a standard curve. Here we use both 25 µM ATP with 20 µM SAMS peptide substrates. Linearity is shown here under initial velocity conditions when raw data is converted to ADP formed. The AMPK enzyme reaction took place for 60 minutes at room temperature. The enzymatic reaction was then halted using Stop & Detect Buffer B included in the kit and assay plates were read.

    Human AMPK Enzyme Titration

    Linear Response

    Screen for Inhibitors & Perform SAR

    Transcreener Assays are designed for screening compound libraries in a high throughput format. Follow-up SAR can also be performed using the assay to determine inhibitor potency with ease. Assay conditions include 0.5 µg/mL AMPK, 25 µM ATP, 20 µM SAMS for an enzyme reaction of 60 minutes at room temperature.

    Dose-Response - Staurosporine

    Staurosporine Dose Response with AMPK

    IC50 = 2.18 nM

    Robust Assay Yields Quality Data

    Z’ measurements using optimized AMPK reaction conditions indicate a robust assay.  Robust data like this is vital for sizeable high throughput screens that are difficult to complete due to massive sample quantity. Both reactions have 25 μM ATP and 20 μM SAMS. Z' shown here = 0.85.

    Three Fluorescent Readouts Available

    Choose the readout that is the best fit for your lab based on preference and plate reader compatibility. 

    Fluorescent Intensity

    Assay conditions for the AMPK titration shown include 25 µM ATP, 20 µM SAMS, and 24 µg/mL ADP² antibody.

    TR-FRET

    Assay conditions for the AMPK titration shown include 25 µM ATP, 20 µM AMPK,  55 nM tracer, and 8 nM ADP² Tb Antibody.

    Far-Red FP, FI & TR-FRET Readouts Validated on Major Multimode Readers

    Supplier Instrument FP Assays
    		 FI Assays
    		 TR-FRET Assays
    bioteklogo TriStar²S LB 942 in review validated validated
    Mithras² LB 943 in review validated validated
    bioteklogo Cytation™ 5 validated validated validated
    Cytation™ 3 validated validated validated
    Cytation™ 1 validated validated validated
    Synergy™ H1 validated validated validated
    Synergy™ 2/H4/4 validated validated validated
    Synergy™ HTX not capable validated not capable
    Synergy™ Neo 2 validated validated validated
    BMGLABTECH Logo POLARstar® Omega validated validated validated
    FLUOstar® Omega not capable validated validated
    PHERAstar® FSX validated validated validated
    PHERAstar® Plus/FS validated validated validated
    CLARIOstar® /Plus validated validated validated
    hidex logo Sense in review validated in review
    MDS AT logo Analyst® GT/HT validated validated validated
    Gemini® XPS/EM not capable validated not capable
    SpectraMax® M2/M2e not capable validated not capable
    SpectraMax® M5/M5e/FlexStation® 3 not capable validated validated
    SpectraMax® Paradigm validated contact us contact us
    SpectraMax® iD3/iD5 in review validated validated
    perkinElmerLogo EnVision®/EnVision® Xcite validated validated validated
    tecanLogo Infinite® M1000/M1000Pro/Safire2™ validated validated validated
    Infinite® M200 not capable validated not capable
    Infinite® F500 validated validated validated
    Infinite® F200/Ultraevolution contact us validated contact us
    Spark™ 10M validated validated validated

    What's Included

    What You Will Need

    Transcreener ADP2 AMPK Assay Kits

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

    Assay Plates for FI & FP

    Cat. No.Product DescriptionPrice 
    2233Corning 96-Well Black Assay Plates (#3686), 3-Pack (200+ Assays)$40.00Add to Cart
    2229Corning 384-Well Black Assay Plates (#4514), 3-Pack (1,000+ Assays)$45.00Add to Cart
    2230Corning 384-Well Black Assay Plates (#4514), 30-Pack (10,000+ Assays)$400.00Add to Cart

    Assay Plates for TR-FRET

    Cat. No.Product DescriptionPrice 
    2234Corning 96-Well White Assay Plates (#3642), 3-Pack (200+ Assays)$40.00Add to Cart
    2231Corning 384-Well White Assay Plates (#4513), 3-Pack (1,000+ Assays)$50.00Add to Cart
    2232Corning 384-Well White Assay Plates (#4513), 30-Pack (10,000+ Assays)$450.00Add to Cart

    AMPK 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 AMPK 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

    Please fill out the form below. We will respond quickly to get the conversation moving and learn how we can help. We keep things discrete, confidential, and professional.

    BellBrook Labs
    5500 Nobel Drive, Suite 230
    Madison, Wisconsin 53711 USA
    (608) 443-2400

    info@bellbrooklabs.com

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