Akt, also known as protein kinase B (PKB), is a central kinase in such diverse cellular
processes as glucose uptake, cell cycle progression, and apoptosis. Three highly
homologous members define the Akt family: Akt1 (PKBa), Akt2 (PKBb), and Akt3
(PKBg). Akt1 is the most ubiquitously expressed family member. All three Akts contain
an amino-terminal pleckstrin homology domain, a central kinase domain, and a carboxyl-
terminal regulatory domain.
Applications:
Suitable for use in ELISA, Western Blot, Immunocytochemistry, Flow Cytometry. Other applications not tested.
Recommended Dilution:
Flow Cytometry: 50ug/ml. This antibody was validated for flow cytometry using MCF-7 cells. For
intracellular staining to detect Akt1, cells must first be fixed and permeabilized using 4% paraformaldehyde and 0.1% saponin in PBS. Dilute this antibody to 50ug/ml and add 10ul of the diluted solution to 1-5x10e5 cells in a total reaction volume not exceeding 20ulL. The binding of unlabeled monoclonal antibodies may be visualized by adding a secondary developing reagent such as goat anti-mouse IgG conjugated to a fluorochrome. Cells should be washed for a final time in 0.1% saponin prior to flow cytometric analysis.
Immunocytochemistry: 8-25ug/ml to detect Akt1 in fixed human peripheral blood mononuclear cells.
Optimal dilutions to be determined by the researcher.
Storage and Stability:
Lyophilized powder may be stored at 4 degrees C for short-term only. Reconstitute to nominal volume by adding sterile 40-50% glycerol and store at -20 degrees C. Reconstituted product is stable for 12 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
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