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  3. PathScan® Cell Growth Multi-Target Sandwich ELISA Kit

PathScan® Cell Growth Multi-Target Sandwich ELISA Kit #7239

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    Product Specifications

    REACTIVITY H M
    Application Key:
    • ELISA+-ELISA and/or ELISA-like Assays 
    Species Cross-Reactivity Key:
    • H-Human 
    • M-Mouse 

    Product Information

    Product Description

    CST’s PathScan® Cell Growth Multi-Target Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that combines the reagents necessary to detect endogenous levels of S6 ribosomal protein, phospho-S6 ribosomal protein (Ser235/236), Akt1, phospho-Akt (Ser473), phospho-Akt (Thr308) and phospho-p44/42 MAPK (Thr202/Tyr204). These molecules represent key signaling proteins in pathways controlling growth and differentiation. Sixteen assays are provided for each target protein. Specific assay formulations for the indicated target proteins can be found in the datasheets associated with the individual sandwich ELISA kits*. Briefly, a capture antibody** has been coated onto the microwells. After incubation with cell lysates, the target protein is captured by the coated antibody. Following extensive washing, a detection antibody** is added to detect the captured target protein. An HRP-linked secondary antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of absorbance for this developed color is proportional to the quantity of bound target protein. *See companion products. **Antibodies in kit are custom formulations specific to kit.

    实验步骤

    Specificity / Sensitivity

    PathScan® Cell Growth Multi-Target Sandwich ELISA Kit #7239 detects endogenous levels of six proteins: S6 ribosomal protein, phospho-S6 ribosomal protein (Ser235/236), Akt1, phospho-Akt (Ser473), phosho-Akt (Thr308) and phospho-p44/42 MAPK (Thr202/Tyr204). Activation of these proteins can be observed over time in response to PDGF. As shown in Figure 1, stimulation of serum-starved NIH/3T3 cells with PDGF promotes phosphorylation of Akt1 at Thr308 and Ser473, S6 ribosomal protein at Ser235/236 and p44/42 MAPK at Thr202/Tyr204. The level of each target protein (phospho and nonphospho) remains unchanged throughout the 80 minute time course as demonstrated by Western analysis. The relationship between the protein concentration of the lysate and the absorbance at 450 nm can be found in the datasheets associated with the individual PathScan® Sandwich ELISA Kits*. *See companion products. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

    Species Reactivity:

    Human, Mouse

    Background

    Akt is a protooncogene with a critical regulatory role in diverse cellular processes including growth, survival and the cell cycle. Akt is also a major regulator of insulin signaling and glucose metabolism (1-4). Akt is activated by PI3 kinase signaling and activation loop phosphorylation at Thr308 by PDK1 and by phosphorylation within the carboxy terminus at Ser473 by the mTOR-rictor complex (TORC1) (5-7).
    Both p44 and p42 MAP kinases (Erk1 and Erk2) function in a protein kinase cascade that plays a critical role in the regulation of cell growth and differentiation (8-13). MAP kinases are activated by a wide variety of extracellular signals including growth and neurotrophic factors, cytokines, hormones and neurotransmitters. Activation of MAP kinases occurs through phosphorylation of threonine and tyrosine (202 and 204 of human MAP kinase or 183 and 185 of rat MAP kinase) at the sequence T*EY* by a single upstream MAP kinase kinase (MEK) (14,15).
    To effectively promote growth and cell division in a sustained manner, growth factors and mitogens must upregulate translation (16,17). Growth factors and mitogens induce the activation of p70 S6 kinase, which in turn phosphorylates the S6 ribosomal protein. Phosphorylation of S6 ribosomal protein correlates with an increase in translation, particularly of mRNAs with an oligopyrimidine tract in their 5' untranslated regions (17). This group of mRNAs (5'TOP) encodes proteins involved in cell cycle progression and proteins that are part of the translational machinery, such as ribosomal proteins and elongation factors (17,18). The main in vivo S6 ribosomal protein phosphorylation sites, including Ser235, Ser236, Ser240 and Ser244, are located within a small 19 amino acid region in the S6 carboxy terminus (19,20).
    1. Kim, D. and Chung, J. (2002) J Biochem Mol Biol 35, 106-15.
    2. Zdychová, J. and Komers, R. (2005) Physiol Res 54, 1-16.
    3. Song, G. et al. J Cell Mol Med 9, 59-71.
    4. Manning, B.D. and Cantley, L.C. (2007) Cell 129, 1261-74.
    5. Alessi, D.R. et al. (1996) EMBO J 15, 6541-51.
    6. Sarbassov, D.D. et al. (2005) Science 307, 1098-101.
    7. Jacinto, E. et al. (2006) Cell 127, 125-37.
    8. McKay, M.M. and Morrison, D.K. (2007) Oncogene 26, 3113-21.
    9. Pearson, G. et al. (2001) Endocr Rev 22, 153-83.
    10. Marshall, C.J. (1995) Cell 80, 179-85.
    11. Hunter, T. (1995) Cell 80, 225-36.
    12. Hill, C.S. and Treisman, R. (1995) Cell 80, 199-211.
    13. Cowley, S. et al. (1994) Cell 77, 841-52.
    14. Sturgill, T.W. et al. (1988) Nature 334, 715-8.
    15. Payne, D.M. et al. (1991) EMBO J 10, 885-92.
    16. Dufner, A. and Thomas, G. (1999) Exp Cell Res 253, 100-9.
    17. Peterson, R.T. and Schreiber, S.L. (1998) Curr Biol 8, R248-50.
    18. Jefferies, H.B. et al. (1997) EMBO J 16, 3693-704.
    19. Ferrari, S. et al. (1991) J Biol Chem 266, 22770-5.
    20. Flotow, H. and Thomas, G. (1992) J Biol Chem 267, 3074-8.

    Alternate Names

    40S ribosomal protein S6; AKT; AKT serine/threonine kinase 1; AKT1; AKT1 kinase; AKT1m; ERK; ERK-1; ERK-2; ERK1; ERK2; ERT1; ERT2; Extracellular signal-regulated kinase 1; Extracellular signal-regulated kinase 2; extracellular signal-regulated kinase-2; extracellular signal-related kinase 1; HS44KDAP; HUMKER1A; Insulin-stimulated MAP2 kinase; MAP kinase 1; MAP kinase 2; MAP kinase 3; MAP kinase isoform p42; MAP kinase isoform p44; MAPK 1; MAPK 2; MAPK 3; MAPK1; MAPK2; MAPK3; MGC20180; MGC99656; Microtubule-associated protein 2 kinase; Mitogen-activated protein kinase 1; Mitogen-activated protein kinase 2; Mitogen-activated protein kinase 3; MK01; MK03; p38; p40; p41; p41mapk; p42-MAPK; P42MAPK; p44-ERK1; p44-MAPK; P44ERK1; P44MAPK; Phosphoprotein NP33; PKB; PKB alpha; PKB-ALPHA; PRKBA; PRKM1; PRKM2; PRKM3; Protein kinase B; Protein kinase B alpha; protein tyrosine kinase ERK2; Proto-oncogene c-Akt; RAC; rac protein kinase alpha; RAC-ALPHA; RAC-alpha serine/threonine-protein kinase; RAC-PK-alpha; ribosomal protein S6; RPS6; RS6; S6; S6 ribosomal protein; serine-threonine protein kinase; Small ribosomal subunit protein eS6; v-akt murine thymoma viral oncogene homolog 1; v-akt murine thymoma viral oncogene-like protein 1

    Database Links

    UniProt ID: P27361 , P62753 , P28482 , P31749


    Entrez-Gene Id: 5595 , 6194 , 5594 , 207


    限制使用

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    For Research Use Only. Not for Use in Diagnostic Procedures.
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