Product Pathways - PathScan ELISA
PathScan® Phospho-TrkA (Tyr674/675) Sandwich ELISA Kit #7212
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|Kit Includes||Volume||Solution Color|
|TrkA mAb coated microwells||96 tests|
|P-TrkA (Y674/675) Detection Ab||11 ml||Green|
|Anti-rabbit IgG, HRP-linked Antibody||11 ml||Red|
|TMB Substrate #7004||11 ml||Colorless|
|STOP Solution #7002||11 ml||Colorless|
|Sealing Tape||2 sheets|
|ELISA Wash Buffer (20X)||25 ml||Colorless|
|ELISA Sample Diluent||25 ml||Blue|
|Cell Lysis Buffer (10X) #9803||15 ml||Yellowish|
Note: 12 8-well modules –Each module is designed to break apart for 8 tests.
Note: Kit should be stored at 4°C with the exception of Cell Lysis Buffer, which is stored at –20°C (packaged separately).
Species enclosed in parentheses are predicted to react based on 100% sequence homology.
CST's PathScan® Phospho-TrkA (Tyr674/675) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects transfected levels of Phospho-TrkA (Tyr674/675) protein. A TrkA Mouse mAb has been coated onto the microwells. After incubation with cell lysates, both phospho- and nonphospho-TrkA proteins are captured by the coated antibody. Following extensive washing, a Phospho-TrkA (Tyr674/675) rabbit antibody is added to detect phospho-TrkA protein. Anti-rabbit IgG, HRP-linked 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-TrkA (Tyr674/675) protein.
Specificity / Sensitivity
CST's PathScan® Phospho-TrkA (Tyr674/675) Sandwich ELISA Kit #7212 detects transfected levels of phospho-TrkA (Tyr674/675) Protein. As shown in Figure 1, using this ELISA Kit #7212, a significant induction of phospho-TrkA (Tyr490) is detected in 3T3/TrkA cells treated with NGF. However, the levels of total TrkA (phospho and nonphospho) in untreated and NGF-treated cells detected by PathScan® Total TrkA Sandwich ELISA Kit #7208, remain unchanged. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.Antibodies in kit are custom formulations specific to kit.
ELISA - Western correlation
Figure 1. Treatment of 3T3/TrkA cells with NGF stimulates phosphorylation of TrkA at Tyr674/675, detected by PathScan® Phospho-TrkA (Tyr674/675) Sandwich ELISA kit #7212, but does not affect the level of total TrkA detected by PathScan® Total TrkA Sandwich ELISA kit #7208. OD 450 readings are shown in the top figure, while the corresponding Western blots using Phospho-TrkA (Tyr674/675) Antibody #4610 (right panel) or TrkA Rabbit mAb #2505 (left panel), are shown in the bottom figure. The human TrkA is expressed in 3T3/TrkA cells.
The family of Trk receptor tyrosine kinases consists of TrkA, TrkB, and TrkC. While the sequence of these family members is highly conserved, they are activated by different neurotrophins: TrkA by NGF, TrkB by BDNF or NT4, and TrkC by NT3 (1). Neurotrophin signaling through these receptors regulates a number of physiological processes, such as cell survival, proliferation, neural development, and axon and dendrite growth and patterning (1). In the adult nervous system, the Trk receptors regulate synaptic strength and plasticity. TrkA regulates proliferation and is important for development and maturation of the nervous system (2). Phosphorylation at Tyr490 is required for Shc association and activation of the Ras-MAP kinase cascade (3,4). Residues Tyr674/675 lie within the catalytic domain, and phosphorylation at these sites reflects TrkA kinase activity (3-6). Point mutations, deletions, and chromosomal rearrangements (chimeras) cause ligand-independent receptor dimerization and activation of TrkA (7-10). TrkA is activated in many malignancies including breast, ovarian, prostate, and thyroid carcinomas (8-13). Research studies suggest that expression of TrkA in neuroblastomas may be a good prognostic marker as TrkA signals growth arrest and differentiation of cells originating from the neural crest (10).
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- Segal, R.A. and Greenberg, M.E. (1996) Annu Rev Neurosci 19, 463-89.
- Stephens, R.M. et al. (1994) Neuron 12, 691-705.
- Marsh, H.N. et al. (2003) J Cell Biol 163, 999-1010.
- Obermeier, A. et al. (1993) EMBO J 12, 933-41.
- Obermeier, A. et al. (1994) EMBO J 13, 1585-90.
- Arevalo, J.C. et al. (2001) Oncogene 20, 1229-34.
- Reuther, G.W. et al. (2000) Mol Cell Biol 20, 8655-66.
- Greco, A. et al. (1997) Genes Chromosomes Cancer 19, 112-23.
- Pierotti, M.A. and Greco, A. (2006) Cancer Lett 232, 90-8.
- Lagadec, C. et al. (2009) Oncogene 28, 1960-70.
- Greco, A. et al. (2010) Mol Cell Endocrinol 321, 44-9.
- Ødegaard, E. et al. (2007) Hum Pathol 38, 140-6.
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- 7210 PathScan® Phospho-TrkA (Tyr490) Sandwich ELISA Kit
- 7208 PathScan® Total TrkA Sandwich ELISA Kit
- 9141 Phospho-TrkA (Tyr490) Antibody
- 4621 Phospho-TrkA (Tyr674/675)/TrkB (Tyr706/707) (C50F3) Rabbit mAb
- 2505 TrkA Antibody
- 2508 TrkA (14G6) Rabbit mAb
- 7074 Anti-rabbit IgG, HRP-linked Antibody
- 7004 TMB Substrate
- 7002 STOP Solution
For Research Use Only. Not For Use In Diagnostic Procedures.