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Human FGFR4/β-Klotho Reporter Assay Kit

SIZE SKU PRICE
1 x-96 well format assays—
3 x-32 assays in-96 well format—
SIZE SKU
3 x-32 assays in-96 well format
1 x-96 well format assays

Product Description and Product Data

This is an all-inclusive cell-based luciferase reporter assay kit targeting the Human Fibroblast Growth Factor Receptor 4 and β-Klotho (FGFR4/β-Klotho). INDIGO’s FGFR4/β-Klotho reporter assay utilizes proprietary mammalian cells that have been engineered to provide constitutive expression of the Human Fibroblast Growth Factor Receptor 4 and β-Klotho. In addition to FGFR4/β-Klotho Reporter Cells, this kit provides two optimized media for use during cell culture and in diluting the user’s test samples, a reference agonist, Luciferase Detection Reagent, and a cell culture-ready assay plate. The principal application of this assay is in the screening of test samples to quantify any functional activity, either agonist or antagonist, that they may exert against human FGFR4/β-Klotho. This kit provides researchers with clear, reproducible results, exceptional cell viability post-thaw, and consistent results lot to lot. Kits must be stored at -80C. Do not store in liquid nitrogen. Note: reporter cells cannot be refrozen or maintained in extended culture.

Features

  • Ready to Use Upon Receipt

  • Includes All Needed Components
  • Contains Transfected Reporter Cells
  • Eliminates Cell Licensing Fees
  • Clear, Reproducible Results
  • Consistent Results Lot to Lot

Product Specifications

Target TypeGrowth Factor Receptor
SpeciesHuman
Receptor FormHybrid
Assay ModeAgonist, Antagonist
Kit Components
  • FGFR4/β-Klotho Reporter Cells
  • Cell Recovery Medium (CRM)
  • Compound Screening Medium (CSM)
  • FGF-19, (ref. activator; in PBS/0.1% BSA )
  • Detection Substrate
  • Detection Buffer
  • White, sterile, cell-culture ready assay plate
Shelf Life6 months
Shipping RequirementsDry Ice
Storage temperature-80C

Data

Activation of FGFR4/β-Klotho. Activation assays were performed using the endocrine reference activator FGF-19 (provided) and FGF-21. Polypeptide ligands were procured from Peprotech. The absence of signal in FGF-19 treated ‘Mock’ cells (which contains the NFAT-Luc reporter vector, but do not express FGFR4 or β-Klotho) confirms that the observed ligand-dependent response is specific to FGFR4/β-Klotho activation.
Inhibition of FGFR4/β-Klotho. FGFR4/β-Klotho cells were treated with an EC80 concentration of the reference activator FGF-19 and varying concentrations of the FGFR inhibitors JNJ-4275493, FIIN-3, FIIN-2, BGJ398 and PD173074 (all compounds obtained from Cayman Chemical). INDIGO’s Live Cell Multiplex (LCM) Assay confirmed that no treatment concentrations were cytotoxic (data not shown). For both Activation and Inhibition assays, luminescence was quantified and values of average (n=3) relative light units (RLU), corresponding standard deviation (SD), Fold-Activation, and Z’ values were calculated. GraphPad Prism software was used to plot data using the leastsquares method of non-linear regression for Fold-Activation or RLU vs. Log10 [Cmpd], and to determine EC50 / IC50 values.

Target Background

The family of Fibroblast Growth Factors (FGFs) comprise approximately 23 members that are related by core sequence and structure conservation, with the majority of FGFs being secreted signaling proteins. Secreted FGFs are predominantly autocrine and paracrine factors, with only three members evolved to function as endocrine factors1. FGFs bind and activate FGF Receptors (FGFRs) which, themselves, are members of the family of high-affinity tyrosine kinase receptors.

Paracrine FGFs show high affinity towards the extracellular matrix (ECM) component heparin sulfate (HS) and are thus retained in the ECM and function locally. In contrast, the atypical endocrine subfamily of FGFs, that comprise FGF-19, FGF-21, and FGF-23, have reduced affinity for HS and can therefore escape from the ECM into the circulation to reach target distant organs. However, this subfamily typically requires association with members of the Klotho family of proteins as cofactors for efficient binding to their cognate receptor(s).

Aberrant FGF-19 signaling mediated by its receptor, FGFR4 and co-receptor β-Klotho, is known to be a driver of hepatocellular carcinoma (HCC). Consequently, FGF-19 and FGFR4/β-Klotho command considerable interest as therapeutic targets in drug development and drug safety screening.

Citations

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