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PathScan^® Phospho-p70 S6 Kinase (Thr389) Sandwich ELISA Kit

PathScan^® Phospho-p70 S6 Kinase (Thr389) Sandwich ELISA Kit #7063

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ELISA

Supporting Data

REACTIVITY

H M

Application Key:

ELISA-ELISA

Species Cross-Reactivity Key:

H-Human
M-Mouse

Product Information

Product Description

The PathScan^® Phospho-p70 S6 Kinase (Thr389) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of phospho-p70 S6 kinase phosphorylated at Thr389. A p70 S6 kinase rabbit mAb has been coated onto the microwells. After incubation with cell lysates, p70 S6 kinase is captured by the coated antibody. Following extensive washing, a phospho-p70 S6 kinase (Thr389) mouse detection mAb is added to detect the captured phospho-p70 S6 kinase (Thr389). Anti-mouse IgG, HRP-linked antibody is then used to recognize the bound detection antibody. The HRP substrate TMB is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of p70 S6 kinase phosphorylated at Thr389.

*Antibodies in this kit are custom formulations specific to kit.

Protocol

Specificity / Sensitivity

The PathScan^® Phospho-p70 S6 Kinase (Thr389) Sandwich ELISA Kit detects endogenous levels of p70 S6 kinase when phosphorylated at Thr389, as shown in Figure 1. This kit is predicted to cross-react with the p85 isoform of S6 kinase. The kit sensitivity is shown in Figure 2. 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

p70 S6 kinase is a mitogen activated Ser/Thr protein kinase that is required for cell growth and G1 cell cycle progression (1,2). p70 S6 kinase phosphorylates the S6 protein of the 40S ribosomal subunit and is involved in translational control of 5' oligopyrimidine tract mRNAs (1). A second isoform, p85 S6 kinase, is derived from the same gene and is identical to p70 S6 kinase except for 23 extra residues at the amino terminus, which encode a nuclear localizing signal (1). Both isoforms lie on a mitogen activated signaling pathway downstream of phosphoinositide-3 kinase (PI-3K) and the target of rapamycin, FRAP/mTOR, a pathway distinct from the Ras/MAP kinase cascade (1). The activity of p70 S6 kinase is controlled by multiple phosphorylation events located within the catalytic, linker and pseudosubstrate domains (1). Phosphorylation of Thr229 in the catalytic domain and Thr389 in the linker domain are most critical for kinase function (1). Phosphorylation of Thr389, however, most closely correlates with p70 kinase activity in vivo (3). Prior phosphorylation of Thr389 is required for the action of phosphoinositide 3-dependent protein kinase 1 (PDK1) on Thr229 (4,5). Phosphorylation of this site is stimulated by growth factors such as insulin, EGF and FGF, as well as by serum and some G-protein-coupled receptor ligands, and is blocked by wortmannin, LY294002 (PI-3K inhibitor) and rapamycin (FRAP/mTOR inhibitor) (1,6,7). Ser411, Thr421 and Ser424 lie within a Ser-Pro-rich region located in the pseudosubstrate region (1). Phosphorylation at these sites is thought to activate p70 S6 kinase via relief of pseudosubstrate suppression (1,2). Another LY294002 and rapamycin sensitive phosphorylation site, Ser371, is an in vitro substrate for mTOR and correlates well with the activity of a partially rapamycin resistant mutant p70 S6 kinase (8).

Database Links

UniProt ID: P23443

Entrez-Gene Id: 6198

Product Citations: 1

Ganapathy, B. et al. (2016) PLoS One 11, e0155456.

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For Research Use Only. Not For Use In Diagnostic Procedures.

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PathScan® Phospho-p70 S6 Kinase (Thr389) Sandwich ELISA Kit #7063