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HTRF Tag-Lite Adenosine A3 Receptor Cells, 200 Assay Points

Tag-lite cells transiently expressing the A3 receptor labeled with Terbium for use in receptor binding applications. The cells are provided ready to use.

For research use only. Not for use in diagnostic procedures.
Part Number: C1TT1A3
Part Number: C1TT1A3
Unit Size: 200 Assay Points
  • A3 receptor is one of the four adenosine receptor family (including A1, A2A, and A2B receptors). A3 receptor is implicated in various pathological conditions, such as cardiac and cerebral ischaemia, and neurodegenerative diseases, as well as inflammatory pathologies including rheumatoid arthritis and asthma. Thus, it is considered as an interesting therapeutic target.

    Cells expressing the A3 receptor are provided pre-labeled with Terbium, and can be used to conduct receptor binding studies on the aforementioned receptor.

  • Assay Points
    200
    Assay Target Type
    Cells
    Assay Technology
    HTRF
    Brand
    Tag-Lite
    Quantity
    1
    Shelf Life
    540.0 Day(s)
    Shipping Conditions
    Shipped in Dry Ice
    Therapeutic Area
    Cardiovascular
    Infectious Diseases
    Inflammation
    Metabolism/Diabetes
    NASH/Fibrosis
    Neuroscience
    Oncology & Inflammation
    Rare Diseases
    Unit Size
    200 Assay Points
  • Assay principle

    Running your A3 receptor binding assay using Tag-lite is as easy as it can get. Simply dispense 10 µL of labeled cells (C1TT1A3) into each well, followed by 5 µL of labeled ligand (L0069GRE) and 5 µL of the compound you wish to test. Like all HTRF assays, Tag-lite assays do not require any washing steps. A diagram of the procedure to be followed is given on the right.

    gpcr-how-it-works-assay-principle-a3-receptor-c1tt1a3
    Saturation binding (KD)

    A saturation binding assay measures total and non-specific binding for increasing concentrations of ligand under equilibrium conditions. To perform the assay, the fluorescent ligand is titrated into a solution containing a fixed amount of labeled cells, and then incubated to equilibrium. The HTRF Ratio obtained from this titration is the total binding.

    gpcr-how-it-works-saturation-binding-kd-c1tt1a3
    how-it-works-adenosine-a3-receptor-labeled-cells
    Competitive binding (KI)

    A competitive binding assay is performed to measure the dissociation constant, Ki. To perform the assay, the compound is titrated into a solution containing a fixed concentration of fluorescent ligand and a fixed amount of cells.

    gpcr-how-it-works-competitive-binding-ki-c1tt1a3
    how-it-works-adenosine-a3-receptor-labeled-cells

     

  • Kd and Ki validation

    Examples of data obtained using Adenosine A3 receptor labeled cells and their matching fluorescent ligand (L0069GRE). XAC (Xanthine amine congener) was used for Ki determination in the competitive binding assay. Results may vary from one HTRF® compatible reader to another.

    assay-validation-adenosine-a3-labeled-cells
    assay-validation-adenosine-a3-labeled-cells
    Compound profiling on Adenosine A1, A2A, A2B and A3 receptors

    Adenosine receptors designated by four subtypes A1, A2A, A2B and A3 were shown to be promising pharmacological targets for the development of therapeutic compounds for various therapeutic areas such as cancer, neurodegenerative disease and COPD/Asthma. Thus, a wide range of compounds, and particularly antagonists have been largely studied.

    The Tag-Lite® binding assay platform offers the complete adenosine receptors subset A1, A2A, A2B and A3 to characterize compounds in term of binding affinity (Ki) and selectivity profile.

    The set of compounds have been profiled on A1, A2A, A2B and A3 receptors using their respective fluorescent ligands L0067RED, L0058RED, L0068RED and L0069GRE.The selectivity profiles of several compounds involved in different therapeutic areas were assessed with this Tag-Lite platform.

    The table below summarizes the resulting Ki obtained for each receptor subtype for the full panel of compounds. The results are confirming the expected selectivity and the range of affinities for these reference compounds.

    Compound Pharmacological class Compounds affinities (Ki - nM) for the different adenosine receptor subtypes
        A1 A2A A2B A3
    XAC Non selective antagonist 12 15 20 50
    Theophylline Non selective antagonist 4000 5000 18000 14000
    DPCPX A1 antagonist 7.6 170 145 800
    Vipadenant A2A antagonist 70 0.9 55 20000
    Ciforadenant (CPI-444) A2A antagonist 150 3.4 660 1600
    Preladenant (SCH 420814) A2A antagonist >20000 5 >20000 >20000
    Istradefylline (KW-6002) A2A antagonist >10000 40 >10000 >10000
    ZM241385 A2A antagonist 260 0.7 40 750
    AZD4635 A2A antagonist 900 15 660 >10000
    MRS1706 A2B inverse agonist 150 325 30 >300
    MRS1220 A3 antagonist 325 37 >500 1.6


     

    assay-validation-adenosine-a3-labeled-cells
    assay-validation-adenosine-a3-labeled-cells

     

    assay-validation-adenosine-a3-labeled-cells

     

    The graphs show the pattern of the dose-response curves obtained for five compounds displaying different selectivity profile

    assay-validation-adenosine-a3-labeled-cells

     

    assay-validation-adenosine-a3-labeled-cells

     

     

Resources

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Guide
HTRF solutions, guide to major applications

This guide provides you an overview of HTRF applications in several therapeutic areas.