Akt Antibody

9272:

Flow , Immunofluorescence , Immunoprecipitation , Western Blotting

Akt Antibody detects endogenous levels of total Akt1, Akt2 and Akt3 proteins. The antibody does not cross-react with related kinases.

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the carboxy-terminal sequence of mouse Akt. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western blot analysis of extracts from CHO cells, transfected with non-targeted (-) or SignalSilence^® Akt siRNA I (+) siRNA, using Akt Antibody #9272 and p42 MAP Kinase (Erk2) Antibody #9108. The Akt antibody confirms silencing of protein expression while the p42 MAP Kinase (Erk2) antibody was used to control for loading and specificity of Akt siRNA (A). Phospho-GSK-3β (Ser9) Antibody #9336 was used to confirm downstream pathway inhibition (B).

Western Blotting

Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence^® Control siRNA (Fluorescein Conjugate) #6201 (-) or SignalSilence^® Akt siRNA I (+), using Akt Antibody #9272 and p42 MAP Kinase (Erk2) Antibody #9108. Akt antibody confirms silencing of Akt expression, while the p42 MAP kinase (Erk2) antibody is used to control for loading and specificity of Akt siRNA.

Western Blotting

Western blot analysis of extracts from NIH/3T3 cells, untreated or PDGF-treated (50 ng/ml) for the indicated times, using Phospho-Akt (Ser473) Antibody #9271 (upper) or Akt Antibody (lower).

Flow Cytometry

Flow cytometric analysis of Jurkat cells, untreated (blue) or LY294002-treated (green), using Akt Antibody compared to a nonspecific negative control antibody (red).

IF-IC

Confocal immunofluorescent images of C2C12 cells showing nuclear and cytoplasmic localization with Akt Antibody (left, red) compared to an isotype control (right). Actin filaments have been labeled with fluorescein phalloidin.

Akt, also referred to as PKB or Rac, plays a critical role in controlling survival and apoptosis (1-3). This protein kinase is activated by insulin and various growth and survival factors to function in a wortmannin-sensitive pathway involving PI3 kinase (2,3). Akt is activated by phospholipid binding and activation loop phosphorylation at Thr308 by PDK1 (4) and by phosphorylation within the carboxy terminus at Ser473. The previously elusive PDK2 responsible for phosphorylation of Akt at Ser473 has been identified as mammalian target of rapamycin (mTOR) in a rapamycin-insensitive complex with rictor and Sin1 (5,6). Akt promotes cell survival by inhibiting apoptosis through phosphorylation and inactivation of several targets, including Bad (7), forkhead transcription factors (8), c-Raf (9), and caspase-9. PTEN phosphatase is a major negative regulator of the PI3 kinase/Akt signaling pathway (10). LY294002 is a specific PI3 kinase inhibitor (11). Another essential Akt function is the regulation of glycogen synthesis through phosphorylation and inactivation of GSK-3α and β (12,13). Akt may also play a role in insulin stimulation of glucose transport (12). In addition to its role in survival and glycogen synthesis, Akt is involved in cell cycle regulation by preventing GSK-3β-mediated phosphorylation and degradation of cyclin D1 (14) and by negatively regulating the cyclin dependent kinase inhibitors p27 Kip (15) and p21 Waf1/CIP1 (16). Akt also plays a critical role in cell growth by directly phosphorylating mTOR in a rapamycin-sensitive complex containing raptor (17). More importantly, Akt phosphorylates and inactivates tuberin (TSC2), an inhibitor of mTOR within the mTOR-raptor complex (18,19).

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2. Burgering, B.M. and Coffer, P.J. (1995) Nature 376, 599-602.
3. Franke, T.F. et al. (1995) Cell 81, 727-36.
4. Alessi, D.R. et al. (1996) EMBO J 15, 6541-51.
5. Sarbassov, D.D. et al. (2005) Science 307, 1098-101.
6. Jacinto, E. et al. (2006) Cell 127, 125-37.
7. Cardone, M.H. et al. (1998) Science 282, 1318-21.
8. Brunet, A. et al. (1999) Cell 96, 857-68.
9. Zimmermann, S. and Moelling, K. (1999) Science 286, 1741-4.
10. Cantley, L.C. and Neel, B.G. (1999) Proc Natl Acad Sci USA 96, 4240-5.
11. Vlahos, C.J. et al. (1994) J Biol Chem 269, 5241-8.
12. Hajduch, E. et al. (2001) FEBS Lett 492, 199-203.
13. Cross, D.A. et al. (1995) Nature 378, 785-9.
14. Diehl, J.A. et al. (1998) Genes Dev 12, 3499-511.
15. Gesbert, F. et al. (2000) J Biol Chem 275, 39223-30.
16. Zhou, B.P. et al. (2001) Nat Cell Biol 3, 245-52.
17. Navé, B.T. et al. (1999) Biochem J 344 Pt 2, 427-31.
18. Inoki, K. et al. (2002) Nat Cell Biol 4, 648-57.
19. Manning, B.D. et al. (2002) Mol Cell 10, 151-62.

• Asselin, E. et al. (2001) XIAP regulates Akt activity and caspase-3-dependent cleavage during cisplatin-induced apoptosis in human ovarian epithelial cancer cells. Cancer Res. 61, 1862-1868. Applications: Western Blot
• Bommakanti, R. K. et al. (2000) Dual regulation of Akt/protein kinase B by heterotrimeric G protein subunits. J. Biol. Chem. 275, 38870-38876. Applications: Western Blot
• Campbell, R. A. et al. (2001) Phosphatidylinositol 3-kinase/Akt-mediated activation of estrogen receptor alpha: a new model for anti-estrogen resistance. J. Biol. Chem. 276, 9817-9824. Applications: Western Blot
• Otero, D. C. et al. (2000) CD19-dependent activation of Akt kinase in B lymphocytes. J. Biol. Chem. 276, 1474-1478. Applications: Western Blotting
• Fukuda, T. et al. (2003) PINCH-1 is an obligate partner of integrin-linked kinase (ILK) functioning in cell shape modulation, motility, and survival. J. Biol. Chem. 278, 51324-51333. Applications: Western Blotting
• Patrucco, E. et al. (2004) PI3Kgamma modulates the cardiac response to chronic pressure overload by distinct kinase-dependent and -independent effects. Cell 118, 375-387. Applications: Western Blotting
• Huang, H. et al. (2001) PTEN Induces Chemosensitivity in PTEN-mutated Prostate Cancer Cells by Suppression of Bcl-2 Expression. J. Biol. Chem. 276, 38830-38836. Applications: Western Blotting
• Meng, F. et al. (2002) Akt Is a Downstream Target of NF-kappaB. J. Biol. Chem. 277, 29674-29680. Applications: Western Blotting
• Zubiaur, M. et al. (2002) CD38 Is Associated with Lipid Rafts and upon Receptor Stimulation Leads to Akt/Protein Kinase B and Erk Activation in the Absence of the CD3-zeta Immune Receptor Tyrosine-based Activation Motifs. J. Biol. Chem. 277, 13-22. Applications: Western Blotting
• Radisavljevic, Z. et al. (2000) Vascular endothelial growth factor up-regulates ICAM-1 expression via the phosphatidylinositol 3 OH-kinase/AKT/nitric oxide pathway and modulates migration of brain microvascular endothelial cells. J. Biol. Chem. 275, 20770-20774. Applications: Western Blot
• Rohani, M.G. et al. (2010) BMC Immunol 11, 53. Applications: Western Blotting
• Chung, Y.W. et al. (2011) J Biol Chem 286, 29681-90. Applications: Western Blotting
• Sykes, S.M. et al. (2011) Cell 146, 697-708. Applications: Western Blotting
• Matsuura, S. et al. (2012) Blood , . Applications: Western Blotting

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• 4691 Akt (pan) (C67E7) Rabbit mAb
• 4060 Phospho-Akt (Ser473) (D9E) XP^® Rabbit mAb
• 2965 Phospho-Akt (Thr308) (C31E5E) Rabbit mAb
• 9901 LY294002 (PI3 Kinase Inhibitor)
• 7071 Phototope^® -HRP Western Blot Detection System, Anti-rabbit IgG, HRP-linked Antibody
• 7074 Anti-rabbit IgG, HRP-linked Antibody
• 7727 Biotinylated Protein Ladder Detection Pack
• 7720 Prestained Protein Marker, Broad Range (Premixed Format)
• 7003 20X LumiGLO^® Reagent and 20X Peroxide
• 9273 Akt Control Cell Extracts
• 9916 Phospho-Akt Pathway Antibody Sampler Kit

For Research Use Only. Not For Use In Diagnostic Procedures.