Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (BSA and Azide Free)
Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (BSA and Azide Free) #80722
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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). Data were generated using the standard formulation of this product.
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). Data were generated using the standard formulation of this product.
Flow cytometric analysis of Jurkat cells, untreated (green) or treated with LY294002 #9901, Wortmannin #9951 and U0126 #9903 (blue), using Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (solid line) compared to a concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed line). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody. Data were generated using the standard formulation of this product.
Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (BSA and Azide Free) 80722
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Filter:
WB
IF
F
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). Data were generated using the standard formulation of this product.
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). Data were generated using the standard formulation of this product.
Flow cytometric analysis of Jurkat cells, untreated (green) or treated with LY294002 #9901, Wortmannin #9951 and U0126 #9903 (blue), using Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (solid line) compared to a concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed line). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody. Data were generated using the standard formulation of this product.
This product is the carrier free version of product #13038. All data were generated using the same antibody clone in the standard formulation which contains BSA and glycerol.
This formulation is ideal for use with technologies requiring specialized or custom antibody labeling, including fluorophores, metals, lanthanides, and oligonucleotides. It is not recommended for ChIP, ChIP-seq, CUT&RUN, or CUT&Tag assays. If you require a carrier free formulation for chromatin profiling, please contact us. Optimal dilutions/concentrations should be determined by the end user.
Formulation
Supplied in 1X PBS, BSA and Azide Free.
For standard formulation of this product see product #13038.
Storage
Store at -20°C. This product will freeze at -20°C so it is recommended to aliquot into single-use vials to avoid multiple freeze/thaw cycles. A slight precipitate may be present and can be dissolved by gently vortexing. This will not interfere with antibody performance.
Specificity / Sensitivity
Phospho-Akt (Thr308) (D25E6) XP® Rabbit mAb (BSA and Azide Free) recognizes endogenous levels of Akt1 protein only when phosphorylated at Thr308. This antibody also recognizes endogenous levels of Akt2 protein when phosphorylated at Thr309 or Akt3 protein when phosphorylated at Thr305.
Species Reactivity:
Human, Mouse, Rat, Monkey
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Thr308 of human Akt1 protein.
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 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 Kip1 (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).