Product Pathways - Motif Antibodies
PhosphoScan® Kit (P-Tyr-100) #7900
| Kit Includes | Quantity |
|---|---|
| 10X Lysis Buffer | 20 milliliters |
| 10X IAP Buffer | 10 milliliters |
| 10X HEPES Buffer | 30 milliliters |
| Urea | 60 grams |
| Vanadate (100mM) (100X) | 300 microliters |
| DTT (1.25M) | 300 microliters |
| Trypsin-TPCK Solution (1mg/ml) (100X) | 650 microliters |
| Phosphotyrosine Mouse mAb (P-Y-100) Beads | 80 microliters |
| Sep-Pak C18 Columns (0.7 ml each) | 5 units |
| ZipTips | 5 units |
| Control Peptides | 1 assays |
*Product is specific to this kit. **Stored at room temperature, packaged separately.
Description
PhosphoScan® Kit (P-Tyr-100) from Cell Signaling Technology (CST) allows for the purification and identification of tyrosine phosphorylation sites in cellular proteins when coupled with LC tandem mass spectrometry using CST's patented PhosphoScan® technology. The assay is based on the specific enrichment of phosphotyrosine-containing peptides using an antibody against phosphotyrosine. Cells are lysed in a urea-containing buffer, and cellular proteins are digested by protease and fractionated by reversed-phase solid-phase extraction. Peptides are then subjected to immunoaffinity purification using Phosphotyrosine Mouse mAb (P-Tyr-100) #9411 coupled to protein G agarose beads*. Overnight incubation ensures high-affinity binding of phosphotyrosine-containing peptides to P-Tyr-100 beads. Unbound peptides are removed through washing, and phosphotyrosine-containing peptides are eluted with dilute acid. Reversed-phase chromatography is performed on microtips to separate phosphopeptides from antibody and to concentrate them for LC tandem mass spectrometry.The kit also contains control peptides from 2x108 Jurkat cells that were treated with pervandate and calyculin A. These control peptides allow the investigator to determine the efficiency of the immunoaffinity purification (IAP) step and the LC-MS/MS analysis (1 assay).*Product is specific to this kit.
License / Use Restriction
- Notice to Non-Profit/Academic Purchasers: The purchase of this kit product includes the limited right and license to use the kit in the practice of any technology or methods described and claimed in U.S. Patent Number 7,198,896 and foreign equivalents, “Immunoaffinity Isolation of Modified Peptides from Complex Mixtures”, Rush et al., Pub. Date March 6, 2003 (and equivalents), provided such kit is not used in research or development projects funded in whole or in part by For-Profit/Corporate sponsors or in high-throughput screening projects resulting in information that will be offered to For-Profit/Corporate entities on a fee-for-access basis (which use(s) of purchased kit product, in any field, require a separate commercial use license from Cell Signaling Technology, Inc. ).
- Notice to For-Profit/Corporate Purchasers: The purchase of this kit product does not include the right or license to use such kit in the practice of any technology or methods described and claimed in the U.S. Patent Number named in (i) above. Use of purchased kit product in such methods, in any field, requires a separate commercial use license from Cell Signaling Technology, Inc.
- Notice to all purchasers: The purchase of any other non-kit product, including motif antibody products, from CST does not include the right or license to use such non-kit product in the practice of any technology or method described and claimed in the U.S. Patent Number named in (i) above. Use of purchased non-kit product in such methods, in any field, requires a separate commercial use license from Cell Signaling Technology, Inc.
For license inquiries, please contact Chris Bunker, Ph.D., Director of New Business Development, Cell Signaling Technology, Inc. (phone: 978-867-2307, e-mail: cbunker@cellsignal.com)
Overview
Background
PhosphoScan® Kits, based on CST's published method (1), isolate large numbers of cellular phosphorylated peptides, allowing the investigator to obtain a global overview of phosphorylation in the cell, covering many classes of proteins without preconceived biases about where tyrosine phosphorylation sites will be found.
Tyrosine phosphorylation plays a key role in cellular signaling (2). In cancer, unregulated tyrosine kinase activity can drive malignancy and tumor formation by generating inappropriate proliferation and survival signals (3). Current phosphoproteomic approaches generally reveal only small numbers of tyrosine phosphorylation sites, in keeping with the low level of phosphotyrosine relative to phosphoserine and phosphothreonine residues (4). PhosphoScan® Kit, based on CST's published method (1,5), isolates large numbers of cellular phosphotyrosine-containing peptides. This allows the investigator to obtain a global overview of tyrosine phosphorylation in the cell, covering many classes of proteins without preconceived biases about where tyrosine phosphorylation sites will be found.
- Rush, J. et al. (2005) Nat Biotechnol 23, 94-101.
- Schlessinger, J. (2000) Cell 103, 211-25.
- Blume-Jensen, P. and Hunter, T. (2001) Nature 411, 355-65.
- Mann, M. et al. (2002) Trends Biotechnol 20, 261-8.
- Rush, J. et al. (2003) U.S. Patent Publication , 20030044848.
Application References
- Guo, A. et al. (2008) Proc Natl Acad Sci U S A 105, 692-7. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Stokes, M.P. et al. (2007) Proc Natl Acad Sci U S A 104, 19855-60. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Rikova, K. et al. (2007) Cell 131, 1190-203. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Gu, T.L. et al. (2007) Blood 110, 323-33. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Gu, T.L. et al. (2007) Leukemia 21, 563-6. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Mercher, T. et al. (2006) Blood 108, 2770-9. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Walters, D.K. et al. (2006) Leuk Res 30, 1097-104. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Griswold, I.J. et al. (2006) Mol Cell Biol 26, 6082-93. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Gu, T.L. et al. (2006) Blood 108, 4202-4. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Goss, V.L. et al. (2006) Blood 107, 4888-97. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Zhang, Y. et al. (2005) Mol Cell Proteomics 4, 1240-50. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
- Rush, J. et al. (2005) Nat Biotechnol 23, 94-101. This article references the use of PhosphoScan® Kit (P-Tyr-100) in the following applications: Phosphoscan
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
