Product Pathways - Tyrosine Kinase / Adaptors
PathScan® Phospho-FGF Receptor 3 (panTyr) Chemiluminescent Sandwich ELISA Kit #64740
|Product Includes||Volume||Solution Color|
|ELISA Sample Diluent||25 ml||Blue|
|Phospho Tyrosine Mouse Detection mAb||1 ea||Green (Lyophilized)|
|Anti-mouse IgG, HRP-linked Antibody (ELISA Formulated)||1 ea||Red (Lyophilized)|
|Detection Antibody Diluent||5.5 ml||Green|
|HRP Diluent||5.5 ml||Red|
|Luminol/Enhancer Solution||3 ml||Colorless|
|ELISA Wash Buffer (20X)||25 ml||Colorless|
|Cell Lysis Buffer (10X) #9803||15 ml||Yellowish|
|FGFR3 Rabbit mAb Coated Microwells||96 tests|
|Sealing Tape||2 sheets|
|Stable Peroxide Buffer||3 ml||Colorless|
Specificity / Sensitivity
PathScan® Phospho-FGF Receptor 3 (panTyr) Chemiluminescent Sandwich ELISA Kit #64740 detects endogenous levels of tyrosine-phosphorylated FGFR3 in human cells. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.
The PathScan® Phospho-FGF Receptor 3 (panTyr) Chemiluminescent Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of tyrosine-phosphorylated FGFR3 protein with a chemiluminescent readout. Chemiluminescent ELISAs often have a wider dynamic range and higher sensitivity than conventional chromogenic detection. This chemiluminescent ELISA, which is offered in low volume microplates, shows increased signal and sensitivity while using a smaller sample size. An FGFR3 rabbit mAb has been coated on the microwells. After incubation with cell lysates, both phospho- and nonphospho-FGFR3 proteins are captured by the coated antibody. Following extensive washing, a phospho tyrosine mouse mAb is added to detect the captured tyrosine-phosphorylated FGFR3 protein. Anti-mouse IgG, HRP-linked antibody is then used to recognize the bound detection antibody. Chemiluminescent reagent is added for signal development. The magnitude of light emission, measured in relative light units (RLU), is proportional to the quantity of tyrosine-phosphorylated FGFR3 protein.
Antibodies in kit are custom formulations specific to kit.
Figure 2. The relationship between protein concentration of phospho or nonphospho lysates and immediate light generation with chemiluminescent substrate as detected by the PathScan® Phospho-FGF Receptor 3 (panTyr) Chemiluminescent Sandwich ELISA Kit is shown. Unstarved KMS-11 cells were cultured (0.8 x 106 cells/ml) and lysed with or without addition of phosphatase inhibitors to the lysis buffer (phospho or nonphospho lysate, respectively).
ELISA - Western correlation
Figure 1. Constitutive tyrosine-phosphorylation of FGFR3 in KMS-11 cells lysed in the presence of phosphatase inhibitors (+) is detected by PathScan® Phospho-FGF Receptor 3 (panTyr) Chemiluminescent Sandwich ELISA Kit #64740. In contrast, a low level of phospho-FGFR3 protein is detected in KMS-11 cells lysed in the absence of phosphatase inhibitors* (-). For the FGFR3-negative RPMI 8226 cells, there is no phospho-FGFR3 detected by this ELISA kit in either presence or absence of phosphatase inhibitors. Immediate light generation with chemiluminescent substrate is shown. *Phosphatase inhibitors include sodium pyrophosphate, β-glycerophosphate, and sodium orthovanadate.
Fibroblast growth factors (FGFs) produce mitogenic and angiogenic effects in target cells by signaling through cell surface receptor tyrosine kinases. There are four members of the FGF receptor family: FGFR1 (flg), FGFR2 (bek, KGFR), FGFR3, and FGFR4. Each receptor contains an extracellular ligand binding domain, a transmembrane domain, and a cytoplasmic kinase domain (1). Following ligand binding and dimerization, the receptors are phosphorylated at specific tyrosine residues (2). Seven tyrosine residues in the cytoplasmic tail of FGFR1 can be phosphorylated: Tyr463, 583, 585, 653, 654, 730, and 766. Tyr653 and Tyr654 are important for catalytic activity of activated FGFR and are essential for signaling (3). The other phosphorylated tyrosine residues may provide docking sites for downstream signaling components such as Crk and PLCγ (4,5).
Activating mutations within fibroblast growth factor receptor 3 (FGFR3) are responsible for human skeletal dysplasias including achondroplasia and the neonatal lethal syndromes thanatophoric dysplasia types I and II (6,7). Several of these same FGFR3 mutations as well as overexpression of FGFR3 proteins have also been identified somatically in human cancers, including multiple myeloma, bladder carcinoma and cervical cancer (8). Thus, FGFR3 may represent a potential target for therapy.
- Powers, C.J. et al. (2000) Endocr Relat Cancer 7, 165-97.
- Reilly, J.F. et al. (2000) J Biol Chem 275, 7771-8.
- Mohammadi, M. et al. (1996) Mol Cell Biol 16, 977-89.
- Mohammadi, M. et al. (1991) Mol Cell Biol 11, 5068-78.
- Larsson, H. et al. (1999) J Biol Chem 274, 25726-34.
- Wilkie, A.O. et al. (2002) Am J Med Genet 112, 266-78.
- Yamashita, A. et al. (2014) Nature 513, 507-11.
- Miyake, M. et al. (2007) Biochem Biophys Res Commun 362, 865-71.
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- 4574 FGF Receptor 3 (C51F2) Rabbit mAb
- 7076 Anti-mouse IgG, HRP-linked Antibody
- 9411 Phospho-Tyrosine Mouse mAb (P-Tyr-100)
- 9803 Cell Lysis Buffer (10X)
- 9808 Phosphate Buffered Saline (PBS-20X)
- 9809 Phosphate Buffered Saline with Tween® 20 (PBST-20X)
- 9998 BSA
For Research Use Only. Not For Use In Diagnostic Procedures.
U.S. Patent No. 5,675,063.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
PathScan is a trademark of Cell Signaling Technology, Inc.
Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.
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