|Reactivity:||Hu, Ms, Rt|
WB 1:1,000 ICC/IF: 1:100 IHC: 1:100
|Purification:||antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues around Thr308 of mouse Akt1|
Background: 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 by phosphorylating and inactivating 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 (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 mTORraptor complex (18). Inhibition of mTOR stops the protein synthesis machinery due to inactivation of its effector, p70 S6 kinase and activation of the eukaryotic initiation factor 4E binding protein 1 (4E-EP1), an inhibitor of translation (18,19). Specificity/Sensitivity: Phospho-Akt (Thr308) (D25E6) Rabbit mAb detects endogenous levels of Akt only when phosphorylated at Thr308.
|Buffer:||PBS with 0.02% sodium azide,50% glycerol,pH7.3.|
|Storage:||Store at -20℃. Avoid freeze / thaw cycles.|
Western blot analysis of extracts of HeLa cells, using Phospho-AKT1-T308 antibody at 1:1000 dilution. HeLa cells were treated by 10% FBS for after serum-starvation overnight. Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) at 1:10000 dilution. Lysates/proteins: 25ug per lane. Blocking buffer: 3% BSA. Detection: ECL West Pico Exposure time: 90s.
Immunofluorescence analysis of HeLa cells using Phospho-AKT1-T308 antibody at dilution of 1:100. Blue: DAPI for nuclear staining.
Experiment Type: Western blot (WB) Sample: rat cells Description: Western blot analysis of extracts of rat cells, using antibody at 1:1000 dilution.
(1) Background References: (2) Franke, T.F. et al. (1997) Cell 88, 435–7. (3) Burgering, B.M. and Coffer, P.J. (1995) Nature 376, 599–602. (4) Franke, T.F. et al. (1995) Cell 81, 727–36. (5) Alessi, D.R. et al. (1996) EMBO J 15, 6541–51. (6) Sarbassov, D.D. et al. (2005) Science 307, 1098–101. (7) Jacinto, E. et al. (2006) Cell 127, 125–37. (8) Cardone, M.H. et al. (1998) Science 282, 1318–21. (9) Brunet, A. et al. (1999) Cell 96, 857–68. (10) Zimmermann, S. and Moelling, K. (1999) Science 286, 1741–4.