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Technical Data Sheet

EK6160

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EGFR 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
EP1931 Jhaveri, T.J. et al. (2015) Oncotarget. 6(17):14754 WB: HCC1806 cells
EP1931 Kamekura, R. et al. (2014) Oncogene. 33(36): 4531. WB: human SK-CO15
RS3251 Kawasaki, H. et al. (2013) World J Gastroenter. 19(17):2629. WB, ICC: mouse intestinal myofibroblasts
EP1871 Choi, S. et al. (2012) Am J Pathol. 180(1):410. WB: mouse skeletal muscle
EP1871 Kolegraff, K. et al. (2011) Mol Biol Cell. 22(8):1121. WB: human intestinal epithelial cells
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 EGFR phospho-regulation antibody sampler kit can be used to detect EGFR phosphorylation at Ser-1142, Ser-967, and Tyr-1101. The kit also includes an antibody to examine total EGFR expression levels, and secondary reagents for rabbit polyclonal and mouse monoclonal antibody detection.
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Western blot image of human A431 cells treated with Calyculin A (100 nM) for 30 min. Blot lanes were untreated (lanes 1, 3, & 5) or treated with lambda phosphatase (lanes 2, 4, & 6) then probed with anti-EGFR (a.a. 961-972) (lanes 1 & 2), anti-EGFR (Ser-967) (lanes 3 & 4), or anti-EGFR (Ser-1142) (lanes 5 & 6).

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Immunocytochemical labeling in A431 cells untreated or treated with EGF (100 ng/ml) for 5 min. The cells were labeled with anti-EGFR or anti-EGFR (Tyr-1101) monoclonal antibodies, then detected using appropriate secondary antibody conjugated to Cy3.

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Background
The epidermal growth factor receptor (EGFR) is a transmembrane glycoprotein with an extracellular ligand-binding domain and a cytoplasmic domain with intrinsic tyrosine kinase activity. The cytoplasmic domain has a C-terminal region with multiple autophosphorylation sites (Tyr-992, 1068, 1086, 1148, and 1173). These sites are important for downstream signaling and rapid internalization. In addition, EGFR activation leads to c-Src mediated phosphorylation of Tyr-845 and Tyr-1101. The former site is required for mitogenic responses to EGFR activation, while the latter may be an SH2 binding site. Phosphorylation of EGFR on serine and threonine residues is thought to represent a mechanism for regulation of receptor kinase activity and internalization. These sites include a PKC site (Thr-654), CAMKII sites (Ser-1046, 1047, 1057, and 1142), and constitutively phosphorylated sites (Ser-967 and Ser-1002). Thus, the regulation of EGFR activity involves a complex series of phosphorylation events at multiple sites throughout the intracellular portion of the receptor.

Background References
Carpenter, G. (2000) Bioessays 22:697.
Boeri Erba, E. et al. (2005) Mol. Cell. Prot. 4:1107.
Morandell, S. et al. (2008) Proteomics. 8(21):4383.
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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