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ELISA Kit Carbohydrate Metabolic Process

Also showing ELISA Kit ELISA Carbohydrate Metabolic Process

$489
96 assays
1 Kit
PathScan® Total Insulin Receptor β Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects transfected levels of Insulin Receptor β protein. An Insulin Receptor β Mouse mAb has been coated on the microwells. After incubation with cell lysates, Insulin Receptor β protein (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, an Insulin Receptor β Rabbit mAb is added to detect captured Insulin Receptor β protein. Anti-rabbit IgG, HRP-linked antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of Insulin Receptor β protein.Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human

Background: Insulin receptor (INSR) is a membrane receptor tyrosine kinase. The receptor molecule consists of a disulfide linked heterodimer. The α subunit is a 135 kDa extracellular fragment, and the β subunit is a 95 kDa fragment containing an extracellular domain, a single transmembrane domain, and an intracellular tyrosine kinase domain (1). Insulin ligand binding to this receptor results in receptor autophosphorylation and tyrosine kinase activation. INSR catalyzes the tyrosine phosphorylation of molecules such as IRS, Gab1, Shc, and Cbl, which further activate the downstream MAPK, PI3K, and TC10 pathways. This eventually leads to increases in glucose uptake and metabolism as well as cell growth (2,3). INSR has peptide substrate specificity similar to other receptor tyrosine kinase members, preferring acidic residues at the -1 to -4 positions and large hydrophobic amino acids at positions +1 and +3 (4).

$489
96 assays
1 Kit
The PathScan® Phospho-Insulin Receptor β (panTyr) Sandwich ELISA Kit from Cell Signaling Technology is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects transfected levels of tyrosine-phosphorylated Insulin Receptor β protein. An Insulin Receptor β Rabbit mAb has been coated on the microwells. After incubation with cell lysates, Insulin Receptor β protein (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, a phospho-tyrosine Mouse mAb is added to detect captured tyrosine-phosphorylated Insulin Receptor β protein. Anti-mouse IgG, HRP-linked antibody is then used to recognize the bound detection antibody. HRP substrate TMB is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of Insulin Receptor β protein phosphorylated on tyrosine.Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human

Background: Insulin receptor (INSR) is a membrane receptor tyrosine kinase. The receptor molecule consists of a disulfide linked heterodimer. The α subunit is a 135 kDa extracellular fragment, and the β subunit is a 95 kDa fragment containing an extracellular domain, a single transmembrane domain, and an intracellular tyrosine kinase domain (1). Insulin ligand binding to this receptor results in receptor autophosphorylation and tyrosine kinase activation. INSR catalyzes the tyrosine phosphorylation of molecules such as IRS, Gab1, Shc, and Cbl, which further activate the downstream MAPK, PI3K, and TC10 pathways. This eventually leads to increases in glucose uptake and metabolism as well as cell growth (2,3). INSR has peptide substrate specificity similar to other receptor tyrosine kinase members, preferring acidic residues at the -1 to -4 positions and large hydrophobic amino acids at positions +1 and +3 (4).

$489
96 assays
1 Kit
CST's PathScan® Phospho-Insulin Receptor β (Tyr1150/1151) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects transfected phospho-insulin receptor (Tyr1150/1151) protein. An Insulin Receptor β Mouse mAb has been coated onto the microwells. After incubation with cell lysates, both phospho- and nonphospho-insulin receptor proteins are captured by the coated antibody. Following extensive washing, Phospho-IGF-I Receptor β (Tyr1135/1136)/Insulin Receptor β (Tyr1150/1151) Rabbit mAb is added to detect the captured phospho-insulin receptor (Tyr1150/1151) protein. Anti-rabbit IgG, HRP-linked Antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of phospho-insulin receptor β (Tyr1150/1151) protein.Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human, Mouse

Background: Type I insulin-like growth factor receptor (IGF-IR) is a transmembrane receptor tyrosine kinase that is widely expressed in many cell lines and cell types within fetal and postnatal tissues (1-3). Receptor autophosphorylation follows binding of the IGF-I and IGF-II ligands. Three tyrosine residues within the kinase domain (Tyr1131, Tyr1135, and Tyr1136) are the earliest major autophosphorylation sites (4). Phosphorylation of these three tyrosine residues is necessary for kinase activation (5,6). Insulin receptors (IRs) share significant structural and functional similarity with IGF-I receptors, including the presence of an equivalent tyrosine cluster (Tyr1146/1150/1151) within the kinase domain activation loop. Tyrosine autophosphorylation of IRs is one of the earliest cellular responses to insulin stimulation (7). Autophosphorylation begins with phosphorylation at Tyr1146 and either Tyr1150 or Tyr1151, while full kinase activation requires triple tyrosine phosphorylation (8).

$489
96 assays
1 Kit
CST's PathScan® Phospho-Insulin Receptor β (Tyr1146) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects transfected phospho-insulin receptor (Tyr1146) protein. A Phospho-Insulin Receptor (Tyr1146) Antibody has been coated onto the microwells. After incubation with cell lysates, phospho-insulin receptor (Tyr1146) proteins are captured by the coated antibody. Following extensive washing, Insulin Receptor β Mouse mAb is added to detect the captured phospho-insulin receptor (Tyr1146) protein. Anti-mouse IgG, HRP-linked Antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of phospho-insulin receptor (Tyr1146) protein.Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human

Background: Type I insulin-like growth factor receptor (IGF-IR) is a transmembrane receptor tyrosine kinase that is widely expressed in many cell lines and cell types within fetal and postnatal tissues (1-3). Receptor autophosphorylation follows binding of the IGF-I and IGF-II ligands. Three tyrosine residues within the kinase domain (Tyr1131, Tyr1135, and Tyr1136) are the earliest major autophosphorylation sites (4). Phosphorylation of these three tyrosine residues is necessary for kinase activation (5,6). Insulin receptors (IRs) share significant structural and functional similarity with IGF-I receptors, including the presence of an equivalent tyrosine cluster (Tyr1146/1150/1151) within the kinase domain activation loop. Tyrosine autophosphorylation of IRs is one of the earliest cellular responses to insulin stimulation (7). Autophosphorylation begins with phosphorylation at Tyr1146 and either Tyr1150 or Tyr1151, while full kinase activation requires triple tyrosine phosphorylation (8).

$489
96 assays
1 Kit
The PathScan® Phospho-Akt (Thr308) Chemiluminescent Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of phospho-Akt (Thr308) protein with a chemiluminescent readout. Chemiluminescent ELISAs often have a wider dynamic range and higher sensitivity than conventional chromogenic detection. This chemiluminescent ELISA which is offered in low volume microplates, shows increased signal and sensitivity while using a smaller sample size. An Akt rabbit antibody has been coated on the microwells. After incubation with cell lysates, both phospho- and nonphospho-Akt proteins are captured by the coated antibody. Following extensive washing, phospho-Akt (Thr308) mouse antibody is added to detect the captured phospho-Akt protein. Anti-mouse IgG, HRP-linked antibody is then used to recognize the bound detection antibody. Chemiluminescent reagent is added for signal development. The magnitude of light emission, measured in relative light units (RLU), is proportional to the quantity of phospho-Akt (Thr308) protein.Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human, Mouse

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).

$489
96 assays
1 Kit
CST's PathScan® Phospho-Akt (Thr308) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects phospho-Akt (Thr308) protein. An Akt Antibody has been coated onto the microwells. After incubation with cell lysates, both phospho- and nonphospho-Akt proteins are captured by the coated antibody. Following extensive washing, Phospho-Akt (Thr308) Mouse mAb is added to detect the captured phospho-Akt protein. Anti-mouse IgG, HRP-linked Antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of phospho-Akt (Thr308) protein.Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human, Mouse

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).

$499
96 assays
1 Kit
The FastScan™ Phospho-Akt (Thr308) ELISA Kit is a sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Akt when phosphorylated at Thr308. To perform the assay, sample is incubated with a capture antibody conjugated with a proprietary tag and a second detection antibody linked to HRP, forming a sandwich with phospho-Akt (Thr308) in solution. This entire complex is immobilized to the plate via an anti-tag antibody. The wells are then washed to remove unbound material. TMB is then added. The magnitude of observed signal is proportional to the quantity of phospho-Akt (Thr308). Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human, Monkey, Mouse, Rat

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).

$499
96 assays
1 Kit
The FastScan™ Phospho-Akt (Ser473) ELISA Kit is a sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Akt when phosphorylated at Ser473. To perform the assay, sample is incubated with a capture antibody conjugated with a proprietary tag and a second detection antibody linked to HRP, forming a sandwich with phospho-Akt (Ser473) in solution. This entire complex is immobilized to the plate via an anti-tag antibody. The wells are then washed to remove unbound material. TMB is then added. The magnitude of observed signal is proportional to the quantity of phospho-Akt (Ser473). Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human, Monkey, Mouse, Rat

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).

$489
96 assays
1 Kit
CST's PathScan® Total Akt2 Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Akt2 protein. An Akt rabbit antibody has been coated on the microwells. After incubation with cell lysates, Akt protein (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, Akt2 mouse mAb is added to detect captured Akt2 protein. HRP-linked anti-mouse IgG is then used to recognize the bound detection antibody. HRP substrate TMB is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of Akt2 protein.
REACTIVITY
Human, Mouse

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).

$489
96 assays
1 Kit
CST's PathScan® Phospho-Akt2 (Ser474) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Akt2 protein when phosphorylated at Ser474. A phospho-Akt rabbit antibody has been coated on the microwells. After incubation with cell lysates, phospho-Akt protein is captured by the coated antibody. Following extensive washing, Akt2 mouse mAb is added to detect captured Akt2 protein. HRP-linked anti-mouse IgG is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of Akt2 phosphorylated at Ser474.Antibodies in kit are custom formulations specific to kit.
REACTIVITY
Human, Mouse

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).

$489
96 assays
1 Kit
CST's PathScan® Total Akt2 Sandwich ELISA Kit (Mouse Preferred) is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Akt2 protein. An Akt rabbit antibody has been coated on the microwells. After incubation with cell lysates, Akt protein (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, Akt2 mouse antibody is added to detect captured Akt2 protein. Anti-mouse IgG, HRP-linked antibody* is then used to recognize the bound detection antibody. HRP substrate TMB is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of Akt2 protein. *Antibodies in this kit are custom formulations specific to the kit.
REACTIVITY
Mouse

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).

$489
96 assays
1 Kit
CST's PathScan® Phospho-Akt2 (Ser474) Sandwich ELISA Kit (Mouse Preferred) is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Akt2 protein phosphorylated at Ser474. A phospho-Akt rabbit antibody has been coated onto the microwells. After incubation with cell lysates, phospho-Akt protein is captured by coated antibody. Following extensive washing, Akt2 mouse antibody is added to detect the captured phospho-Akt2 protein. Anti-mouse IgG, HRP-linked antibody* is then used to recognize the bound detection antibody. HRP substrate TMB is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of Akt2 phosphorylated at Ser474. *Antibodies in this kit are custom formulations specific to the kit.
REACTIVITY
Mouse

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).