Revision 1
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP

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:

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UniProt ID:

#Q16539

Entrez-Gene Id:

1432

Product Information

Storage

Store at -20°C

Specificity / Sensitivity

Species Reactivity:

Human, Mouse, Rat

Product Description

Nonradioactive p38 MAP Kinase Assay Kit provides all the reagents necessary to measure p38 MAP kinase activity in cells. First, Immobilized Phospho-p38 MAPK (Thr180/Tyr182) mAb is used to immunoprecipitate p38 MAP kinase, then an in vitro kinase assay is performed using ATF-2 as a substrate. ATF-2 phosphorylation is detected by Western blotting using Phospho-ATF-2 (Thr71) Antibody.

Background

p38 MAP kinase (MAPK), also called RK (1) or CSBP (2), is the mammalian orthologue of the yeast HOG kinase that participates in a signaling cascade controlling cellular responses to cytokines and stress (1-4). Four isoforms of p38 MAPK, p38α, β, γ (also known as Erk6 or SAPK3), and δ (also known as SAPK4) have been identified. Similar to the SAPK/JNK pathway, p38 MAPK is activated by a variety of cellular stresses, including osmotic shock, inflammatory cytokines, lipopolysaccharide (LPS), UV light, and growth factors (1-5). MKK3, MKK6, and SEK activate p38 MAPK by phosphorylation at Thr180 and Tyr182. Activated p38 MAPK has been shown to phosphorylate and activate MAPKAP kinase 2 (3) and to phosphorylate the transcription factors ATF-2 (5), Max (6), and MEF2 (5-8). SB203580 (4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-imidazole) is a selective inhibitor of p38 MAPK. This compound inhibits the activation of MAPKAPK-2 by p38 MAPK and subsequent phosphorylation of HSP27 (9). SB203580 inhibits p38 MAPK catalytic activity by binding to the ATP-binding pocket, but does not inhibit phosphorylation of p38 MAPK by upstream kinases (10).

  1. Rouse, J. et al. (1994) Cell 78, 1027-37.
  2. Han, J. et al. (1994) Science 265, 808-11.
  3. Lee, J.C. et al. (1994) Nature 372, 739-46.
  4. Freshney, N.W. et al. (1994) Cell 78, 1039-49.
  5. Raingeaud, J. et al. (1995) J Biol Chem 270, 7420-6.
  6. Zervos, A.S. et al. (1995) Proc Natl Acad Sci U S A 92, 10531-4.
  7. Zhao, M. et al. (1999) Mol Cell Biol 19, 21-30.
  8. Yang, S.H. et al. (1999) Mol Cell Biol 19, 4028-38.
  9. Cuenda, A. et al. (1995) FEBS Lett 364, 229-33.
  10. Kumar, S. et al. (1999) Biochem Biophys Res Commun 263, 825-31.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation

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

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

限制使用

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Revision 1
#9820

p38 MAP Kinase Assay Kit (Nonradioactive)

Western Blotting Image 1: p38 MAP Kinase Assay Kit (Nonradioactive) Expand Image
使用 Phospho-ATF-2 (Thr71) Antibody 对经过紫外线处理的 NIH-3T3 细胞的 p38 MAP 激酶活性进行蛋白质印迹分析。细胞提取物与固定的 p38 MAPK (Thr180/Tyr182) mAb 一起孵育过夜。在 100 μM 的冷 ATP 和 1 μg 的 ATF-2 融合蛋白的存在下进行激酶反应。使用 Phospho-ATF-2 (Thr71) Antibody 进行蛋白质印迹实验来检测 ATF-2 在 Thr71 位点的 磷酸化。
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