Revision 4
#9106
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For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:
W, W-S, FC-FP
Reactivity:
H M R Hm Mk Mi Z B Pg
Sensitivity:
Endogenous
MW (kDa):
42, 44
Source/Isotype:
Mouse IgG1
UniProt ID:
#P27361, #P28482
Entrez-Gene Id:
5595, 5594
Product Usage Information
Application | Dilution |
---|---|
Western Blotting | 1:2000 |
Simple Western™ | 1:10 - 1:50 |
Flow Cytometry (Fixed/Permeabilized) | 1:1000 - 1:2000 |
Storage
Specificity/Sensitivity
Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (E10) Mouse mAb detects endogenous levels of p44 and p42 MAP Kinase (Erk1 and Erk2) when dually phosphorylated at Thr202 and Tyr204 of Erk1 (Thr185 and Tyr187 of Erk2), but not singly phosphorylated at Thr202 or Tyr204. This antibody does not cross-react with the corresponding phosphorylated residues of either SAPK/JNK or p38 MAP kinase. This antibody may cross-react with an unknown cytoskeletal protein in some cell lines as visualized by immunofluorescence.
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr202/Tyr204 of human p44 MAP kinase.
Background
Mitogen-activated protein kinases (MAPKs) are a widely conserved family of serine/threonine protein kinases involved in many cellular programs, such as cell proliferation, differentiation, motility, and death. The p44/42 MAPK (Erk1/2) signaling pathway can be activated in response to a diverse range of extracellular stimuli, including mitogens, growth factors, and cytokines (1-3), and research investigators consider it an important target in the diagnosis and treatment of cancer (4). Upon stimulation, a sequential three-part protein kinase cascade is initiated, consisting of a MAP kinase kinase kinase (MAPKKK or MAP3K), a MAP kinase kinase (MAPKK or MAP2K), and a MAP kinase (MAPK). Multiple p44/42 MAP3Ks have been identified, including members of the Raf family, as well as Mos and Tpl2/COT. MEK1 and MEK2 are the primary MAPKKs in this pathway (5,6). MEK1 and MEK2 activate p44 and p42 through phosphorylation of activation loop residues Thr202/Tyr204 and Thr185/Tyr187, respectively. Several downstream targets of p44/42 have been identified, including p90RSK (7) and the transcription factor Elk-1 (8,9). p44/42 are negatively regulated by a family of dual-specificity (Thr/Tyr) MAPK phosphatases, known as DUSPs or MKPs (10), along with MEK inhibitors, such as U0126 and PD98059.
Background References
- Roux, P.P. and Blenis, J. (2004) Microbiol Mol Biol Rev 68, 320-44.
- Baccarini, M. (2005) FEBS Lett 579, 3271-7.
- Meloche, S. and Pouysségur, J. (2007) Oncogene 26, 3227-39.
- Roberts, P.J. and Der, C.J. (2007) Oncogene 26, 3291-310.
- Rubinfeld, H. and Seger, R. (2005) Mol Biotechnol 31, 151-74.
- Murphy, L.O. and Blenis, J. (2006) Trends Biochem Sci 31, 268-75.
- Dalby, K.N. et al. (1998) J Biol Chem 273, 1496-505.
- Marais, R. et al. (1993) Cell 73, 381-93.
- Kortenjann, M. et al. (1994) Mol Cell Biol 14, 4815-24.
- Owens, D.M. and Keyse, S.M. (2007) Oncogene 26, 3203-13.
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 nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
Applications Key
W: Western Blotting 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
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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.
Revision 4
Specificity and sensitivity of the Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (E10) Mouse mAb: This antibody reacts specifically with as little as 50 pg of phosphorylated MAP kinase and does not cross-react with up to 4 µg of nonphosphorylated MAP kinase by Western blotting.
Western blot analysis of extracts from FGF treated SK-N-MC cells, using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (E10) Mouse mAb (upper) or control p44/42 MAPK (Erk1/2) Antibody #9102 (lower).
Simple Western™ analysis of lysates (0.1 mg/mL) from serum-starved HeLa cells treated with TPA (200 nM, 15 min) using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (E10) Mouse mAb #9106. The virtual lane view (left) shows the target bands (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
Revision 4
Flow cytometric analysis of Jurkat cells, treated with U0126 #9903 (10 uM, 2 hr; blue) or treated with TPA (12-O-Tetradecanoylphorbol-13-Acetate #4174 (200 nM, 30 min; green) using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (E10) Mouse mAb (solid lines) or concentration-matched Mouse (G3A1) mAb IgG1 Isotype Control #5415 (dashed lines). Anti-mouse IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4408 was used as a secondary antibody.