Human c-MET/HGFR Reporter Assay Kit

SIZE	SKU	PRICE
1 x-384 well format assays	IB30302	—
1 x-96 well format assays	IB30301	—

SIZE	SKU
1 x-384 well format assays	IB30302
1 x-96 well format assays	IB30301

Specifications
Data
Target Background
Documentation
Overview
Citation
Related Products

Product Description and Product Data

This is an all-inclusive cell-based luciferase reporter assay kit targeting the the Human c-MET / Hepatocyte Growth Factor Receptor (c-MET/HGFR). INDIGO’s c-MET/HGFR reporter assay utilizes proprietary mammalian cells that have been engineered to provide constitutive expression of the OXTR. In addition to c-MET/HGFR 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 c-MET/HGFR. 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

Clear, Reproducible Results
All-Inclusive Assay Systems
Exceptional Cell Viability Post-Thaw
Consistent Results Lot to Lot

Product Specifications

Target Type		Growth Factor Receptor
Species		Human
Receptor Form		Hybrid
Assay Mode		Agonist, Antagonist
Kit Components		Human c-MET/HGFR Reporter Cells Cell Recovery Medium (CRM) Compound Screening Medium (CSM) HGF, (ref. agonist; in PBS/0.1%BSA) Detection Substrate Detection Buffer White, sterile, cell-culture ready assay plate
Shelf Life		6 months
Shipping Requirements		Dry Ice
Storage temperature		-80C

Data

Activation of c-MET. Activation assays were performed according to the protocol provided in the Technical Manual using the reference activator HGF (provided). Luminescence was quantified and values of average (n = 3) relative light units (RLU), corresponding standard deviation (SD), Fold-Activation, and Z’ values were calculated. Non-linear regression analyses of Fold-Activation or RLU vs. Log10 [Compound, nM] and EC50 / IC50 values were determined using GraphPad Prism software.

Inhibition of c-MET. c-MET reporter cells were co-treated with an EC80 concentration of the reference activator HGF and varying concentrations of the c-MET inhibitors INCB-28060, SGX523, PHA-665752 and SU11274 (all procured from Cayman Chemical, Ann Arbor MI, USA.) INDIGO’s Live Cell Multiplex (LCM) Assay confirmed that no treatment concentrations were cytotoxic (data not shown). Luminescence was quantified and values of average (n = 3) relative light units (RLU), corresponding standard deviation (SD), Fold-Activation, and Z’ values were calculated. Non-linear regression analyses of Fold-Activation or RLU vs. Log10 [Compound, nM] and EC50 / IC50 values were determined using GraphPad Prism software.

Target Background

The MET proto-oncogene encodes the receptor tyrosine kinase (RTK) c-MET, a.k.a Hepatocyte Growth Factor Receptor (HGFR). The c-MET receptor is formed by proteolytic processing of its precursor protein in the post-Golgi compartment into a singlepass, disulfide-linked α/β heterodimer. This cell surface receptor is expressed in cells of many organs, including the liver, pancreas, prostate, kidney, muscle, and bone marrow.

The only known ligand for c-MET is Hepatocyte Growth Factor (HGF). HGF acts as a pleiotropic factor and cytokine, promoting cell proliferation, survival, motility, differentiation and morphogenesis. The mature form of HGF consists of an α- and β- chain, which are held together by a disulfide bond. HGF binding to c-MET results in receptor homodimerization and phosphorylation of two tyrosine residues located in the intracellular tyrosine kinase domain.

At present, many studies have implicated c-MET in the regulation of cancer cell growth, angiogenesis, invasion and metastasis3. Deregulation and the consequent aberrant signaling of c-MET may occur by different mechanisms including gene amplification, overexpression, activating mutations, and increased ligand-mediated paracrine and autocrine stimulation. It has been established that c-MET is overexpressed in a variety of cancers including Lung, breast, ovary, kidney, colon, thyroid, liver, and gastric carcinomas. Consequently, c-MET and its ligand HGF continue to command much interest as targets for drug development and drug safety screening.