| Product Includes | Product # | Quantity | Color |
|---|---|---|---|
| FastScan™ ELISA Microwell Strip Plate, 96 Well | 53257 | 96 tests | |
| p70 S6 Kinase Rabbit Capture mAb | 77217 | 1 ea | Green (Lyophilized) |
| Phospho-p70 S6 Kinase (Thr389) Mouse HRP-linked mAb | 17081 | 1 ea | Red (Lyophilized) |
| FastScan™ ELISA Capture Antibody Diluent | 16076 | 3 ml | Green |
| FastScan™ ELISA HRP Antibody Diluent | 28120 | 3 ml | |
| TMB Substrate | 7004 | 11 ml | |
| STOP Solution | 7002 | 11 ml | |
| Sealing Tape | 54503 | 1 ea | |
| ELISA Wash Buffer (20X) | 9801 | 25 ml | |
| FastScan™ ELISA Cell Extraction Buffer (5X) | 69905 | 10 ml | |
| FastScan™ ELISA Cell Extraction Enhancer Solution (50X) | 25243 | 1 ml | |
| FastScan™ ELISA Kit #54164 Positive Control Type 1 | 23521 | 1 ea |
*The microwell plate is supplied as 12 8-well modules - Each module is designed to break apart for 8 tests.
Description
The FastScan™ Phospho-p70 S6 Kinase (Thr389) ELISA Kit is a sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of phospho-p70 S6 Kinase (Thr389). 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-p70 S6 Kinase (Thr389) 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-p70 S6 Kinase (Thr389).
*Antibodies in this kit are custom formulations specific to kit.
Specificity/Sensitivity
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).
- Pullen, N. and Thomas, G. (1997) FEBS Lett 410, 78-82.
- Dufner, A. and Thomas, G. (1999) Exp Cell Res 253, 100-9.
- Weng, Q.P. et al. (1998) J Biol Chem 273, 16621-9.
- Pullen, N. et al. (1998) Science 279, 707-10.
- Alessi, D.R. et al. (1998) Curr Biol 8, 69-81.
- Polakiewicz, R.D. et al. (1998) J Biol Chem 273, 23534-41.
- Fingar, D.C. et al. (2002) Genes Dev 16, 1472-87.
- Saitoh, M. et al. (2002) J Biol Chem 277, 20104-12.
Background References
Cross-Reactivity Key
H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected
Trademarks and Patents
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