Revision 6
#2717
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For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:
W, IF-IC, FC-FP
Reactivity:
H M
Sensitivity:
Endogenous
MW (kDa):
70, 72
Source/Isotype:
Rabbit IgG
UniProt ID:
#P43403, #P43405
Entrez-Gene Id:
7535, 6850
Product Usage Information
Application | Dilution |
---|---|
Western Blotting | 1:1000 |
Immunofluorescence (Immunocytochemistry) | 1:200 - 1:800 |
Flow Cytometry (Fixed/Permeabilized) | 1:400 - 1:1600 |
Storage
For a carrier free (BSA and azide free) version of this product see product #31754.
Specificity/Sensitivity
Phospho-Zap-70 (Tyr319)/Syk (Tyr352) (65E4) Rabbit mAb detects endogenous levels of Zap-70 only when phosphorylated at Tyr319. It cross-reacts with endogenous levels of Syk when phosphorylated at Tyr352.
Species predicted to react based on 100% sequence homology
Rat, Hamster, Chicken, Bovine, Dog, Pig, Horse
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Tyr319 of human Zap-70.
Background
The Syk family protein tyrosine kinase Zap-70 is expressed in T and NK cells and plays a critical role in mediating T cell activation in response to T cell receptor (TCR) engagement (1). Following TCR engagement, Zap-70 is rapidly phosphorylated on several tyrosine residues through autophosphorylation and transphosphorylation by the Src family tyrosine kinase Lck (2-6). Tyrosine phosphorylation correlates with increased Zap-70 kinase activity and downstream signaling events. Expression of Zap-70 is correlated with disease progression and survival in patients with chronic lymphocytic leukemia (7,8).
Background References
- Chu, D.H. et al. (1998) Immunol Rev 165, 167-80.
- Iwashima, M. et al. (1994) Science 263, 1136-9.
- Neumeister, E.N. et al. (1995) Mol Cell Biol 15, 3171-8.
- Chan, A.C. et al. (1995) EMBO J 14, 2499-508.
- Williams, B.L. et al. (1999) EMBO J 18, 1832-44.
- Di Bartolo, V. et al. (1999) J Biol Chem 274, 6285-94.
- Wiestner, A. et al. (2003) Blood 101, 4944-51.
- Crespo, M. et al. (2003) N Engl J Med 348, 1764-75.
Species Reactivity
Species reactivity is determined by testing in at least one approved application (e.g., western blot).
Western Blot Buffer
IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
Applications Key
W: Western Blotting IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized)
Cross-Reactivity Key
H: Human M: Mouse R: Rat Hm: Hamster Mk: Monkey Vir: Virus Mi: Mink C: Chicken Dm: D. melanogaster X: Xenopus Z: Zebrafish B: Bovine Dg: Dog Pg: Pig Sc: S. cerevisiae Ce: C. elegans Hr: Horse GP: Guinea Pig Rab: Rabbit G: Goat All: All Species Expected
Trademarks and Patents
Cell Signaling Technology is a 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
Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.
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.
Revision 6
Western blot analysis of extracts from Jurkat cells treated with hydrogen peroxide (2mM for 2 minutes) or with lambda phosphatase and extracts from Ramos cells treated with anti-human IgM (12 micrograms/ml for 2 minutes) using Phospho-Zap-70 (Tyr319)/Syk (Tyr352) (65E4) Rabbit mAb.
Confocal immunofluorescent analysis of Ramos cells, human IgM treated (left) or untreated (right), using Phospho-Zap-70 (Tyr319)/Syk (Tyr352) (65E4) Rabbit mAb (green). Red = Propidium Iodide (PI)/RNase Staining Solution #4087.
Flow cytometric analysis of Ramos cells, untreated (blue) or treated with anti-IgM (12 µg/mL, 2 min; green), using Phospho-Zap-70 (Tyr319)/Syk (Tyr352) (65E4) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.