This sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Phospho-VEGFR-2 (Tyr1175) protein. A VEGFR-2 Mouse Monoclonal Antibody has been coated onto the microwells. After incubation with cell lysates, both nonphospho-and phospho- VEGFR-2 proteins are captured by the coated antibody. Following extensive washing, a phospho-VEGFR-2 Rabbit Monoclonal Antibody is added to detect the captured phospho-VEGFR-2 protein. HRP-linked anti-rabbit antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of phospho- VEGFR-2 protein. * Antibodies in this kit are custom formulations specific to the kit.
Vascular endothelial growth factor receptor 2 (VEGFR-2, KDR, Flk-1) is a major receptor transducing VEGF-induced signaling in endothelial cells. Upon ligand binding, VEGFR-2 undergoes autophosphorylation and becomes activated (1). Major autophosphorylation sites of VEGFR-2 are located in the kinase insert domain (Tyr951/996) and in the tyrosine kinase catalytic domain (Tyr1054/1059) (2). Activation of the receptor leads to rapid recruitment of adaptor proteins, including Shc, GRB2, PI-3 kinase, Nck and the protein tyrosine phosphatases SHP-1 and SHP-2 (3). The phosphorylation of Tyr1212 provides a docking site for Grb2 binding, and phospho-Tyr1175 binds with the p85 subunit of PI-3 kinase and PLCr, as well as Shb (5, 6). The signaling of VEGFR-2 is necessary for the execution of VEGFstimulated proliferation, chemotaxis and sprouting, as well as survival of cultured endothelial cells in vitro and angiogenesis in vivo (4).
Storage and Stability:Kit should be stored at 4°C wtih the exception of 10X Cell Lysis Buffer, which is stored at -20°C (packaged separately). Stable for 6 months For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
