Revision 5

#7108

Store at +4C

PathScan® Phospho-TrkB (panTyr) Sandwich ELISA Kit

1 Kit

(96 assays)

Species Cross Reactivity:
H

UniProt ID:
#Q16620

Entrez-Gene Id:
#4915

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Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

Web:

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Product Includes Product # Quantity Color Storage Temp
TrkB Mouse mAb Coated Microwells6339696 tests+4C
Phospho-Tyrosine Mouse Detection mAb (Biotinylated)140551 eaGreen (Lyophilized)+4C
HRP-Linked Streptavidin (ELISA Formulated)118051 eaRed (Lyophilized)+4C
Detection Antibody Diluent1333911 mlGreen+4C
HRP Diluent1351511 mlRed+4C
TMB Substrate700411 ml+4C
STOP Solution700211 ml+4C
Sealing Tape545032 ea+4C
ELISA Wash Buffer (20X)980125 ml+4C
ELISA Sample Diluent1108325 mlBlue+4C
Cell Lysis Buffer (10X)980315 ml-20C

Kit contents scale proportionally with size, except sealing tape.
Example: The V1 kit contains 5X the listed quantities above, but will exclude the sealing tape.

For the “C” and “V” kits, the supplied 96-well strip plate consists of twelve 8-well strips in a support frame. This enables custom plate configurations.

Description

The PathScan® Phospho-TrkB (panTyr) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects transfected levels of TrkB when phosphorylated on tyrosine residues. A TrkB mouse antibody has been coated onto the microwells. After incubation with cell lysates, TrkB (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, a biotinylated phospho-tyrosine detection antibody is added to detect tyrosine phosphorylation of the captured TrkB protein. HRP-linked streptavidin is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of TrkB phosphorylated on tyrosines.

*Antibodies in this kit are custom formulations specific to kit.

Specificity/Sensitivity

PathScan® Phospho-TrkB (panTyr) Sandwich ELISA Kit #7108 detects transfected levels of phospho-TrkB protein when tyrosines are phosphorylated(see Figure 1). This kit sensitivity is shown in figure 2. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

Background

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).
The phosphorylation sites are conserved between TrkA and TrkB: Tyr490 of TrkA corresponds to Tyr512 in TrkB, and Tyr674/675 of TrkA to Tyr706/707 in TrkB of the human sequence (14). TrkB is overexpressed in tumors, such as neuroblastoma, prostate adenocarcinoma, and pancreatic ductal adenocarcinoma (15). Research studies have shown that in neuroblastomas, overexpression of TrkB correlates with an unfavorable disease outcome when autocrine loops signaling tumor survival are potentiated by additional overexpression of brain-derived neurotrophic factor (BDNF) (16-18). An alternatively spliced truncated TrkB isoform lacking the kinase domain is overexpressed in Wilms’ tumors and this isoform may act as a dominant-negative regulator of TrkB signaling (17).

Background References

  1. Huang, E.J. and Reichardt, L.F. (2003) Annu Rev Biochem 72, 609-42.
  2. Segal, R.A. and Greenberg, M.E. (1996) Annu Rev Neurosci 19, 463-89.
  3. Stephens, R.M. et al. (1994) Neuron 12, 691-705.
  4. Marsh, H.N. et al. (2003) J Cell Biol 163, 999-1010.
  5. Obermeier, A. et al. (1993) EMBO J 12, 933-41.
  6. Obermeier, A. et al. (1994) EMBO J 13, 1585-90.
  7. Arevalo, J.C. et al. (2001) Oncogene 20, 1229-34.
  8. Reuther, G.W. et al. (2000) Mol Cell Biol 20, 8655-66.
  9. Greco, A. et al. (1997) Genes Chromosomes Cancer 19, 112-23.
  10. Pierotti, M.A. and Greco, A. (2006) Cancer Lett 232, 90-8.
  11. Lagadec, C. et al. (2009) Oncogene 28, 1960-70.
  12. Greco, A. et al. (2010) Mol Cell Endocrinol 321, 44-9.
  13. Ødegaard, E. et al. (2007) Hum Pathol 38, 140-6.
  14. Huang, E.J. and Reichardt, L.F. (2003) Annu. Rev. Biochem. 72, 609-642.
  15. Geiger, T.R. and Peeper, D.S. (2005) Cancer Res 65, 7033-6.
  16. Han, L. et al. (2007) Med Hypotheses 68, 407-9.
  17. Aoyama, M. et al. (2001) Cancer Lett 164, 51-60.
  18. Desmet, C.J. and Peeper, D.S. (2006) Cell Mol Life Sci 63, 755-9.

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

PathScan is a registered trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

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Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 5

#7108

PathScan® Phospho-TrkB (panTyr) Sandwich ELISA Kit

Cell Signaling Technology Logo
Figure 1: Treatment of 3T3/TrkB cells with BDNF (#3897) stimulates tyrosine-phosphorylation of TrkB, detected by PathScan® Phospho-TrkB (panTyr) Sandwich ELISA Kit #7108, but does not affect the level of total TrkB detected by sandwich ELISA. Absorbance at 450 nm is shown in the top figure, while the corresponding Western blots using Phospho-TrkA (Tyr674/675)/TrkB (Tyr706/707) (C50F3) Rabbit mAb #4621 (right panel) or TrkB (80E3) Rabbit mAb #4603 (left panel), are shown in the bottom figure. The human TrkB is transfected and expressed in NIH/3T3 cells.
ELISA Image 1: PathScan<sup>®</sup> Phospho-TrkB (panTyr) Sandwich ELISA Kit
Figure 2: The relationship between protein concentration of lysates from untreated and BDNF-treated 3T3/TrkB cells and the absorbance at 450 nm is shown. After starvation, 3T3/TrkB cells (85% confluence) were treated with BDNF (100 ng/ml) for 2 min at 37ºC then lysed.
ELISA Image 2: PathScan<sup>®</sup> Phospho-TrkB (panTyr) Sandwich ELISA Kit
Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.