Description
Simple VEGF Bioassay, Consistent Results
Vascular endothelial growth factor (VEGF) is an important signaling protein that is secreted from epithelial cells, tumor cells and macrophages. It has many functions, including stimulation of angiogenesis, increase of vascular permeability, enhancement of tumor invasion and survival, and inhibition of antitumor response in Treg cells. There are several VEGF receptor subtypes—VEGFR1, VEGFR2 and VEGFR3. VEGFR2 (also known as KDR) mediates almost all of the known receptor cellular responses to VEGF. VEGF occurs in four isoforms, including VEGF-121, VEGF-165, VEGF-189 and VEGF-206, of which VEGF-121 and -165 are diffusible forms. VEGF-165 is the predominant isoform in the body.
All members of the VEGF family stimulate cellular response by binding to receptors of the receptor tyrosine kinase, namely VEGFR-1 (Flt-1) and VEGFR-2 (Flk-1/KDR). When VEGF binds to KDR, the receptor dimerizes and becomes activated through transphosphorylation.
The VEGF Bioassay is a bioluminescent cell-based assay that measures VEGF stimulation and inhibition of KDR (VEGFR-2) using luciferase as a readout. This assay overcomes many of the limitations of current endothelial cell proliferation assays and can be used for discovery and development of novel biologic therapies aimed at either inducing or inhibiting the VEGF response.
The VEGF Responsive Cells have been engineered to express the response element (RE) upstream of luc2P , as well as exogenous VEGF receptor. When VEGF binds to VEGF Responsive Cells, the receptor transduces intracellular signals resulting in luminescence. The bioluminescent signal is detected and quantified using Bio-Glo™ Luciferase Assay System (Cat.# G7940) and a standard luminometer such as the GloMax® Discover System (Cat.# GM3000).
Recombinant VEGF may be used as a positive control ligand in the VEGF Bioassay (Cat.# GA2001, GA2005). Human VEGF-165, amino acids Ala27–Arg191 (Accession# AAM03108), has a predicted molecular mass of 19kDa and is supplied in a solution of 5mM citric acid, 5mM NaHPO 4 , 0.15M NaCl (pH 4.0) and at a concentration of 200μg/ml.
