KD-Validated Phospho-AKT1 (S124) Rabbit mAb (20 μl)

Background

The gene AKT1 encodes one of the three members of the human AKT serine-threonine protein kinase family which are often referred to as protein kinase B alpha, beta, and gamma. These highly similar AKT proteins all have an N-terminal pleckstrin homology domain, a serine/threonine-specific kinase domain and a C-terminal regulatory domain. These proteins are phosphorylated by phosphoinositide 3-kinase (PI3K). AKT/PI3K forms a key component of many signalling pathways that involve the binding of membrane-bound ligands such as receptor tyrosine kinases, G-protein coupled receptors, and integrin-linked kinase. These AKT proteins therefore regulate a wide variety of cellular functions including cell proliferation, survival, metabolism, and angiogenesis in both normal and malignant cells. AKT proteins are recruited to the cell membrane by phosphatidylinositol 3,4,5-trisphosphate (PIP3) after phosphorylation of phosphatidylinositol 4,5-bisphosphate (PIP2) by PI3K. Subsequent phosphorylation of both threonine residue 308 and serine residue 473 is required for full activation of the AKT1 protein encoded by this gene. Phosphorylation of additional residues also occurs, for example, in response to insulin growth factor-1 and epidermal growth factor. Protein phosphatases act as negative regulators of AKT proteins by dephosphorylating AKT or PIP3. The PI3K/AKT signalling pathway is crucial for tumor cell survival. Survival factors can suppress apoptosis in a transcription-independent manner by activating AKT1 which then phosphorylates and inactivates components of the apoptotic machinery. AKT proteins also participate in the mammalian target of rapamycin (mTOR) signalling pathway which controls the assembly of the eukaryotic translation initiation factor 4F (eIF4E) complex and this pathway, in addition to responding to extracellular signals from growth factors and cytokines, is disregulated in many cancers. Mutations in this gene are associated with multiple types of cancer and excessive tissue growth including Proteus syndrome and Cowden syndrome 6, and breast, colorectal, and ovarian cancers. Multiple alternatively spliced transcript variants have been found for this gene.

Images

	Western blotting analysis using anti-Phospho-AKT1 (S124) antibody (Cat#62471). Total cell lysates (30 μg) from various cell lines were loaded and separated by SDS-PAGE. The blot was incubated with anti-Phospho-AKT1 (S124) antibody (Cat#62471, 1:5,000) and HRP-conjugated goat anti-rabbit secondary antibody (Cat#201, 1:20,000) respectively. Image was developed using FeQ™ ECL Substrate Kit (Cat#226).
	Western blotting analysis using anti-phospho-AKT1 (S124) antibody (Cat#62471). Phospho-AKT1 (S124) expression in wild-type (WT) and AKT1 shRNA knockdown (KD) HeLa cells with 20 μg of total cell lysates. Hsp90 α serves as a loading control. The blot was incubated with anti-phospho-AKT1 (S124) antibody (Cat#62471, 1:5,000) and HRP-conjugated goat anti-rabbit secondary antibody (Cat#201, 1:20,000) respectively. Image was developed using NaQ™ ECL Substrate Kit (Cat#716).
	Flow cytometric analysis of Phospho-AKT1 (S124) expression in C2C12 cells using Phospho-AKT1 (S124) antibody (Cat#62471, 1:2,000). Green, isotype control; red, Phospho-AKT1 (S124).
	Immunocytochemical staining of C2C12 cells with Phospho-AKT1 (S124) antibody (Cat#62471, 1:1,000). Nuclei were stained blue with DAPI; Phospho-AKT1 (S124) was stained magenta with Alexa Fluor® 647. Images were taken using Leica stellaris 5. Protein abundance based on laser Intensity and smart gain: Medium. Scale bar: 20 μm.

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