		Technical Data
A1124-90A	Akt, phosphorylated (Ser473) (Protein kinase B, RAC-alpha serine/threonine-protein kinase, murine thymoma viral (v-akt) oncogene homolog-1, rac protein kinase alpha, v-akt murine thymoma viral oncogene homolog 1)

Description:
Akt/PKB is a Ser/Thr kinase and a major known effecter of the PI3 Kinase pathway. It is involved in multiple signaling pathways that relate to many biological processes including glucose metabolism, cell survival/apoptosis, cell cycle control, angiogenesis, differentiation and cell growth and proliferation. In mammals three isoforms of Akt (Akt1/PKB alpha, Akt2/PKB beta, and Akt3/PKB gamma) exist. They exhibit a high degree of homology, but differ slightly in the localization of their regulatory phosphorylation sites. Akt1 is the predominant isoform that is in most tissues and is thought to have a dominant role in growth, survival, embryonic development and adipocyte differentiation. Akt2 is correlated with the regulation of glucose homeostasis and is the predominant isoform expressed in insulin- responsive tissues. Akt3 is abundant in brain tissue. Each Akt isoform is composed of three functionally distinct regions: an N-terminal Pleckstrin Homology (PH) domain that provides a lipid-binding module to direct Akt to PIP3 at the cell membrane that is necessary for its activation, a central catalytic domain containing Thr308, and a C-terminal hydrophobic motif containing Ser473. The activation of Akt is dependent on a dual regulatory mechanism that requires both its translocation to the plasma membrane and dual phosphorylation on Thr308 and Ser473 by PDK1 and the TORC2 complex, respectively.

Applications:
Suitable for use in Immunofluorescence, Flow Cytometry, ELISA, Western Blot and Immunohistochemistry. Other applications not tested.

Recommended Dilution:
Immunofluorescence: 2ug/ml
Flow Cytometry: 1:800
Western Blot: 0.5-1ug/ml detected phosphorylated AKT in lysates from mouse NIH-3T3 fibroblasts treated with 100ng/ml PDGF for 20 minutes.
Immunohistochemistry: 1:200; Pretreatment with citrate, pH 6 antigen retrieval.
Optimal dilutions to be determined by the researcher.

Storage and Stability:
May be stored at 4°C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20°C. Aliquots are stable for at least 12 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.

Type	Isotype	Clone	Grade
Mab	IgG1,k	9i145	Affinity Purified

Size		Storage	Shipping	Source	Host
100ug		-20°C	Blue Ice	Human	Mouse

Concentration:

~1mg/ml

Immunogen:

Peptide encompassing and including phosphorylated Ser473 of human Akt1.

Purity:

Purified by Protein G affinity chromatography.

Form

Supplied as a liquid in 0.1M Tris-glycine, pH 7.4, 150mM sodium chloride, 0.05% sodium azide.

Specificity:

Recognizes human Akt at ~60kD only when phosphorylated on Ser473. Does not crossreact with Akt1/PKBalpha when it is not phosphorylated on Ser473. Species Crossreactivity: mouse and rat. Species sequence homology: Most vertebrates.

Intended for research use only. Not for use in human, therapeutic, or diagnostic applications.

1. Cohen, P, et al. (1997) FEBS Lett., 410: 3-10 (1997). 2. James, S R, et al. (1996) Biochem. J., 315 ( Pt 3): 709-13 (1996). 3. Alessi, D R, et al. (1997) Curr. Biol., 7: 776-89 (1997). 4. Cross, D A, et al. (1995) Nature, 378: 785-9 (1995). 5. Alessi, D R, et al. (1998) Curr. Biol., 8: 69-81 (1998)