| 货号 | 9916T |
| 描述 | The Phospho-Akt Pathway Antibody Sampler Kit provides an economical means to evaluate the activation status of the Akt signaling pathway, including PTEN and phosphorylated Akt, GSK-3beta, c-Raf and PDK1. The kit includes enough primary and secondary antibodies to perform four Western blot experiments, as well as a specific inhibitor of PI3 kinase (LY294002). |
| 目标/特异性 | Each phospho-specific antibody recognizes the phosphorylated form of its target. Akt Antibody recognizes total Akt protein, independent of its phosphorylation state. |
| 供应商 | CST |
| 背景 | 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). |
| 存放说明 | -20C |
| 参考文献 | Franke, T.F. et al. (1997) Cell 88, 435-7. Burgering, B.M. and Coffer, P.J. (1995) Nature 376, 599-602. Franke, T.F. et al. (1995) Cell 81, 727-36. Alessi, D.R. et al. (1996) EMBO J 15, 6541-51. Sarbassov, D.D. et al. (2005) Science 307, 1098-101. Jacinto, E. et al. (2006) Cell 127, 125-37. Cardone, M.H. et al. (1998) Science 282, 1318-21. Brunet, A. et al. (1999) Cell 96, 857-68. Zimmermann, S. and Moelling, K. (1999) Science 286, 1741-4. Cantley, L.C. and Neel, B.G. (1999) Proc Natl Acad Sci USA 96, 4240-5. Vlahos, C.J. et al. (1994) J Biol Chem 269, 5241-8. Hajduch, E. et al. (2001) FEBS Lett 492, 199-203. Cross, D.A. et al. (1995) Nature 378, 785-9. Diehl, J.A. et al. (1998) Genes Dev 12, 3499-511. Gesbert, F. et al. (2000) J Biol Chem 275, 39223-30. Zhou, B.P. et al. (2001) Nat Cell Biol 3, 245-52. Navé, B.T. et al. (1999) Biochem J 344 Pt 2, 427-31. Inoki, K. et al. (2002) Nat Cell Biol 4, 648-57. Manning, B.D. et al. (2002) Mol Cell 10, 151-62. |
Immunoprecipitation of phospho-Akt (Thr308) from Jurkat cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using Phospho-Akt (Thr308) (D25E6) XP®Rabbit mAb. | |
Western blot analysis of extracts from NIH/3T3 cells, untreated (-) or treated with Human Platelet-Derived Growth Factor AA (hPDGF-AA) #8913 (100 ng/ml, 5 min; +), and untreated (-) LNCaP and PC-3 cells, using Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (upper) or Akt (pan) (C67E7) Rabbit mAb #4691 (lower). | |
Confocal immunofluorescent analysis of C2C12 cells, insulin-treated (100 nM, 15 min; left) or treated with LY294002 #9901 (50 μM, 2 hr; right), using Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye). | |
Western blot analysis of extracts from GSK-3α (-/-) (lanes 1,2) and GSK-3β (-/-) (lanes 3,4) mouse embryonic fibroblast (MEF) cells, λ phosphatase or PDGF-treated, using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb #5558 (upper) and α/β-Tubulin Antibody #2148 (lower). (MEF wild type, GSK-3α (-/-) and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada). | |
Flow cytometric analysis of serum-starved NIH/3T3 cells, untreated (blue) or treated with Human Platelet-Derived Growth Factor AA (hPDGF-AA) #8913 (100 ng/ml, 15 min; green), using Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb. Anti-rabbit IgG (H+L), F(ab)2 Fragment (Alexa Fluor® 647 Conjugate) #4414 was used as a secondary antibody. | |
Western blot analysis of extracts from PC3 cells, HCT116 wild-type and HCT116 PDK1 -/- cells using Phospho-PDK1 (Ser241) (C49H2) Rabbit mAb (upper) and Akt (pan) (C67E7) Rabbit mAb #4691 (lower). (HCT116 wild-type and HCT116 PDK1 -/- cells were kindly provided by Dr. Bert Vogelstein, Johns Hopkins University, Baltimore, MD). | |
Western blot analysis of extracts from PC-3 cells, untreated or LY294002/wortmannin-treated, using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb (upper) or GSK-3β (27C10) Rabbit mAb #9315 (lower). | |
Flow cytometric analysis of NIH/3T3 cells, untreated (blue) or PDGF-treated (green), using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb. | |
Western blot analysis of extracts from GSK-3α (-/-) (lanes 1,2) and GSK-3β (-/-) (lanes 3,4) mouse embryonic fibroblast (MEF) cells, λ phosphatase or PDGF-treated, using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb (upper) and α/β-Tubulin Antibody #2148 (lower). (MEF wild type, GSK-3α (-/-) and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada). | |
Confocal immunofluorescent analysis of wild type mouse embryonic fibroblasts (MEFs) (top row), GSK-3β (-/-) MEFs (middle row) , or PC-3 cells (bottom row), untreated (left), LY294002- and Wortmannin-treated (#9901 and #9951 respectively; center) or lambda phosphatase-treated (right), using Phospho-GSK-3β (Ser9) (D85E12) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye). (MEF wild type and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada). | |
Western blot analysis of extracts from PC3 cells, untreated or λ phosphatase-treated, using Phospho-PDK1 (Ser241) (C49H2) Rabbit mAb #3438 (upper), PDK1 Antibody #3062 (middle) or Akt Antibody #9272 (lower). Western blot分析PC3细胞提取物，未处理组或λ 磷酸酶处理组，所用抗体为Phospho-PDK1 (Ser241) (C49H2) Rabbit mAb #3438兔单抗 (上), PDK1 Antibody #3062 (中) 或 Akt Antibody #9272 (下). | |
Western blot analysis of extracts from wild type (lanes 1,2), GSK-3α (-/-) (lanes 3,4) and GSK-3β (-/-) (lanes 5,6) mouse embryonic fibroblast cells (MEF), untreated or PDGF treated, using Phospho-GSK-3β (Ser9) (5B3) Rabbit mAb #9323 (upper) and GSK-3α/β Antibody (lower). (MEF wild type, GSK-3α (-/-) and GSK-3β (-/-) cells were kindly provided by Dr. Jim Woodgett, University of Toronto, Canada). Western blot分析wild type (泳道 1,2), GSK-3α (-/-) (泳道 3,4) 和 GSK-3β (-/-) (泳道 5,6) 小鼠胚胎成纤维细胞(MEF), 未处理组或 PDGF 处理组，所用抗体为Phospho-GSK-3β (Ser9) (5B3) Rabbit mAb 兔单抗#9323 (上) 和 GSK-3α/β Antibody (下） (MEF wild type, GSK-3α (-/-) and GSK-3β (-/-) 细胞是由加拿大多伦多大学 by Dr. Jim Woodgett友情赠送。 | |
Western blot analysis of extracts from PC3 cells, untreated or λ phosphatase-treated, using Phospho-PDK1 (Ser241) (C49H2) Rabbit mAb (upper), PDK1 Antibody #3062 (middle) or Akt Antibody #9272 (lower). | |
Immunohistochemical analysis of frozen SKOV3 xenograft using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb. | |
Immunohistochemical analysis of paraffin-embedded human breast carcinoma comparing SignalStain® Antibody Diluent #8112 (left) to TBST/5% normal goat serum (right) using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060. |
