Akt is the major known effector of the PI3 Kinase pathway. Generation of PIP3 results in the activation of PDK1, which phosphorylates Akt on Thr308, and another kinase complex, thought to be the mTORC2 complex (or possibly an intermediate kinase linking the two) that phosphorylates Akt on Ser473. These phosphorylations additively activate Akt Ser/Thr kinase activity, and the use of phosphorylation state-specific antibodies directed against either of these sites can imply Akt activation. Activation of Akt can be measured directly by immunoprecipitation followed by phosphorylation of a known substrate with radiolabeled ATP. Akt has been shown to phosphorylate over 70 substrates including TSC1&2 in the mTOR pathway, Bad on Ser136, GSK3 (which is inactivated by this phosphorylation), and the FoxO family of transcription factors (PRAS40, AS160).
Applications:
Suitable for use in Western Blot and Immunoprecipitation.. Other applications not tested.
Recommended Dilution:
Western Blot: 1:500
Optimal dilutions to be determined by the researcher.
Positive Control:
L6 cell lysate, HepG2 lysate.
Storage and Stability:
May be stored at 4 degrees C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20 degrees 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.
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