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Technical Data Sheet

EK6440

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ERK1/2 Phospho-Regulation
Antibody Sampler Kit
Price

$395

Kit Components
Applications: WB = Western blot, E = ELISA, ICC = Immunocytochemistry, IP = Immunoprecipitation, IHC = Immunohistochemistry Species: H = Human, R = Rat, M = Mouse, C = Chicken, F = Fish

Product References
EM2331 Kyjacova, L. et al. (2015) Cell Death Differ. 22(6):898. WB: human DU145
EP4101 H. Huang, et al. (2015) Cardiovasc Res. 108(1):50. WB: mouse heart
EM2061 Park, K. et al. (2013) Mol Cell Biol. 33(4):752. WB: human keratinocytes
EP4101 Lu, J. et al. (2013) Basic Res Cardiol. 108(2):326. WB fluorescence: mouse heart
RS3251 Kawasaki, H. et al. (2013) World J Gastroenter. 19(17):2629. WB, ICC: mouse intestinal myofibroblasts
MS3001 Estrada-Bernal, A. et al. (2011) J Neurooncol. 102:353. Western Blot
RS3251 Estrada-Bernal, A. et al. (2011) J Neurooncol. 102:353. Western Blot
Kit Summary
The ERK phospho-regulation antibody sampler kit can be used to examine ERK1 phosphorylation at Thr-202/Tyr-204 and Thr-207. These phosphorylation sites are also conserved in ERK2. A mouse monoclonal antibody to ERK1 and a rabbit polyclonal antibody to ERK2 are also included for examining total ERK expression levels.
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Western blot analysis of human A431 epithelial cells treated with 100 nM calyculin A for 30 min. (lanes 1, 3, 5, & 7) then the blot was treated with lambda phosphatase (lanes 2, 4, 6, & 8). The blots were probed with polyclonal anti-ERK2 (a.a. 181-195) (lanes 1 & 2), anti-ERK2 (Thr-188) (lanes 3 & 4), anti-ERK1/2 (Thr-202/Tyr-204) (lanes 5 & 6), or monoclonal anti-ERK1 (C-terminal region) (lanes 7 & 8).

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Immunocytochemical labeling of phosphorylated ERK1 in paraformaldehyde-fixed and NP-40-permeabilized rat A7r5 cells treated with calyculin A. The fixed cells were labeled with mouse monoclonal antibodies to anti-ERK1 (EM2331) and anti-ERK1/2 (Thr-202/Tyr-204) (EM2061). The antibodies were detected using Goat anti-Mouse secondary antibodies conjugated to DyLight® 488 (left) and DyLight® 594 (right).

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Background
The ERK1/2 (p44/42) MAPK signaling pathway can be activated in response to a diverse range of extracellular stimuli including mitogens, growth factors, and cytokines. Upon stimulation, a sequential three-part MAP kinase cascade is initiated, consisting of a MAP kinase kinase kinase (MAPKKK), a MAP kinase kinase (MAPKK), and a MAP kinase (MAPK). Activation of the MAPKs, ERK1 and ERK2, leads to phosphorylation of activation loop residues Thr-202/Tyr-204 and Thr-185/Tyr-187, respectively. In addition to dual phosphorylation, ERK1 and 2 are autophosphorylated on Thr-207 or Thr-188, respectively. This phosphorylation is required for nuclear translocation of ERK, and leads to phosphorylation of several nuclear proteins involved in cardiac hypertrophy. Mouse models with mutation of Thr-188 in ERK2 show that this site is critical for ERK-mediated cardiac hypertrophy. Thus, phosphorylation of Thr-188 in ERK2 may be important for controlling the nuclear functions of activated ERK.

Background References
Roux, P.P. & Blenis, J. (2004) Microbiol Mol Biol Rev 68:320.
Murphy, L.O. & Blenis, J. (2006) Trends Biochem Sci 31:268.
Owens, D.M. & Keyse, S.M. (2007) Oncogene 26:3203.
Lorenz, K. et al. (2009) Nat Med. 15(1):75.
Buffer and Storage
Mouse monoclonal and rabbit polyclonal antibodies are supplied in phosphate-buffered saline, 50% glycerol, 1 mg/ml BSA, and 0.05% sodium azide. The secondary reagents are supplied in the same buffer without azide. Store all at –20°C. Stable for 1 year.
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