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Human FGFR4 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 (FGFR4). INDIGO’s FGFR4 reporter assay utilizes proprietary mammalian cells that have been engineered to provide constitutive expression of the Human Fibroblast Growth Factor Receptor 4. In addition to FGFR4 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. 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 Reporter Cells
  • Cell Recovery Medium (CRM)
  • Compound Screening Medium (CSM)
  • FGF-2, (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. Activation assays were performed using the paracrine reference activators FGF-2 (aka FGF-Basic; provided) and FGF-1 (aka FGF-Acidic). The limited activity by the endocrine activator FGF-19, and absence of activity with FGF-21, and FGF-23 demonstrates assay specificity. All polypeptide ligands were procured from Peprotech. 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 least-squares method of non-linear regression for Fold-Activation or RLU vs. Log10 [Cmpd], and to determine EC50 / IC50 values.
Inhibition of FGFR4. FGFR4 cells were treated with an EC80 concentration of the reference activator FGF-2 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 least-squares 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 factors.

FGFs bind and activate four FGF Receptors (FGFR1-4) which, themselves, are members of the family of high-affinity tyrosine kinase receptors. Heparin and heparin sulfate proteoglycans (HSPGs) are essential cofactors for paracrine FGF (e.g., FGF-1 and FGF-2) interactions with FGFRs. The association between paracrine FGFs and HSPGs ensures their limited diffusion and enhanced FGFR binding specificity.

In contrast to the paracrine ligand activators of FGFR, endocrine FGFs (e.g., FGF-19, FGF-21, and FGF-23) have minimal affinity to heparin. Rather, they typically require association with members of the Klotho family of proteins as cofactors for efficient binding to their cognate receptor(s). Although, FGF-23 activation of FGFR3 can occur in a Klotho independent manner, signaling through the PLCγ/calcineurin/NFAT pathway.

The FGFs are broad-spectrum mitogens that, through their receptor interactions, regulate a variety of cellular functions including migration, proliferation, differentiation, metabolism and survival. In particular, FGF/FGFR signaling plays a critical role in regulating metabolism in the kidney, liver, brain, intestine and adipose tissues. Not surprisingly, dysfunctional FGFR signaling can lead to a range of physiological disorders. For example, mutation, amplification, and gene fusion may result in abnormal morphogenesis and the progression of several types of cancer. Consequently, the FGF receptors continue to command much interest as targets for drug development and drug safety screening.

Citations

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