Cytochrome P450 2C9 (CYP2C9) Inhibitor Screening Kit (Fluorometric) (BN01070)

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BN01070
  • Cytochrome P450 2C9 (CYP2C9) Inhibitor Screening Kit (Fluorometric) (BN01070)
  • Cytochrome P450 2C9 (CYP2C9) Inhibitor Screening Kit (Fluorometric) (BN01070)
  • Cytochrome P450 2C9 (CYP2C9) Inhibitor Screening Kit (Fluorometric) (BN01070)
€565

Description

ELISA Kit Technical ManualMSDS

Cytochrome P450 2C9 (CYP2C9) Inhibitor Screening Kit (Fluorometric)

Cytochrome P450 2C9 (CYP2C9, EC 1.14.14.1) is a member of the cytochrome P450 monooxidase (CYP) family of microsomal xenobiotic metabolism enzymes. CYPs are membrane-bound hemeproteins responsible for Phase I biotransformation reactions, in which lipophilic drugs and other xenobiotic compounds are converted to more hydrophilic products to facilitate excretion from the body. CYP2C9 is primarily expressed in liver and catalyzes oxidation of small, weakly acidic or hydrophobic molecules containing an aromatic moiety and at least one hydrogen bond donor. Isoforms of the CYP2C subfamily are responsible for metabolism of nearly 20% of all small molecule drugs commonly used by humans. The human CYP2C9 gene is highly polymorphic and CYP2C9 single nucleotide polymorphisms (SNPs) have been implicated in clinical drug/drug interactions involving widely-prescribed drugs with narrow therapeutic indices such as warfarin, phenytoin and indomethacin. In addition, for drugs whose pharmacological activity requires metabolism from a pro-drug form (such as the antihypertensive drug losartan), CYP2C9 inhibition or allelic deficiency can lead to decreased drug efficacy. Assay Genie's CYP2C9 Inhibitor Screening Kit enables rapid screening of drugs and other new chemical entities (NCEs) for compound-CYP2C9 interaction in a reliable, high-throughput fluorescence-based assay. The kit provides a yeast microsomal preparation of human CYP2C9 and human cytochrome P450 reductase (CPR) enzymes. The assay utilizes a non-fluorescent CYP2C9 substrate that is converted into a highly fluorescent metabolite detected in the visible range (Ex/Em = 415/502 nm), ensuring a high signal-to-background ratio with little interference by autofluorescence. The kit contains a complete set of reagents sufficient for performing 100 reactions in a 96-well plate format.

Figure: (a) 7-hydroxy-4-(trifluoromethyl)coumarin (7-HFC) standard curve. One mole of 7-HFC corresponds to the metabolism of one mole of CYP2C9 substrate. (b) Reaction kinetics of recombinant human CYP2C9 enzyme at 37°C in the presence and absence of the indicated CYP2C9 inhibitors (the solvent control reaction contained assay buffer with 0.3% acetonitrile). (c) Dose-response curves for various CYP2C9 ligands of differing structural and mechanistic classes: the canonical competitive CYP2C9 inhibitor sulfaphenazole, the mechanism-based irreversible inhibitor tienilic acid, the antidepressant fluvoxamine (a competitive inhibitor of several CYP isoforms) and the CYP2C9-selective substrate diclofenac. For dose-response curves, percent activity was calculated for each concentration of
inhibitor by comparison to activity of reactions containing no inhibitor. For each CYP2C9 inhibitor, IC 50 values were derived by 4- parameter logistic curve fitting with each point representing the mean ± SEM of at least three replicates. Assays were performed according to the kit protocol.

Key Information Description

Product SKU

BN01070

Size

100 assays

Detection Method

Fluorescence (Ex/Em 415/502 nm)

Species Reactivity

Eukaryotes

Applications

  • Rapid, high-throughput screening of drugs and novel ligands.
  • Development of structure-activity relationship (SAR) models to predict CYP2C9 inhibition liability of novel compounds.
  • Prediction of adverse drug-drug interaction potential and bioavailability for compounds metabolized by CYP2C9.

Features and Benefits

  • Simple, convenient, highly sensitive
  • Fluorescent assay enables easy determination of CYP1A2 activity in a variety of biological samples
  • The substrate shows low background and a high signal-to-noise ratio
  • Kit includes CYP1A2 inhibitor (naphthoflavone) and a stable, recombinant human CYP1A2 co-expressed with NADPH
  • Reductase as a positive control

Kit Components

  • CYP2C9 Assay Buffer
  • 7-HFC Standard
  • CYP2C9 Inhibitor (Tienilic Acid)
  • NADPH Generating System (100X)
  • Beta-NADP+ Stock (100X)
  • CYP2C9 Substrate
  • Recombinant Human CYP2C9

Storage Conditions

-20°C

Shipping Conditions

Gel Pack

USAGE

For Research Use Only! Not For Use in Humans.

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