PathScan? Multiplex Western Cocktail III: Phospho-Stat1, Phospho-SAPK/JNK, Phospho-S6 Ribosomal Prot

• 产品编号：CST-5303S      品牌：CST       原厂货号：5303S
• 产品分类：抗体 > 一抗 > 磷酸化抗体
• 应用分类：

描述：

该cocktail 中的每个磷酸化抗体都能识别内源性特异靶点磷酸化的蛋白水平。Pin1 antibody能检测它的总靶蛋白水平，同时该抗体还能用于蛋白负载对照。该抗体通过使用合成肽段免疫动物而获得。该抗体经蛋白A和肽亲和层析纯化。PathScan® Multiplex Western Cocktail III为分析一种胞膜上各种蛋白的抑制提供了独特的方法，不用去除免疫印迹膜上的封闭液和抗体，然后再次重新标记。该方法为使用者节约了时间，且调高了准确度，减少了试剂的浪费。该系统让使用者能即时检测phospho-Stat1, phospho-SAPK/JNK和phospho-S6、phospho-HSP27核糖体蛋白。cocktail也包括用于控制蛋白负载的Pin1 Antibody。应答许多配体后，Stat1就被激活。Stat1蛋白似乎对 IFN-alpha、IFN-gamma的应答性十分重要 (1-3)。Stat1蛋白酪氨酸（701位点）的磷酸化诱导了Stat1蛋白的二聚化、细胞核内的移位和DNA的结合(4)。人们发现许多肿瘤中的Stat1蛋白被异常活化(5)。应激活化蛋白激酶/Jun-末端激酶SAPK/JNK更有效、优先地被各种环境应激活化，包括UV、gamma辐射、神经酰胺、和炎性细胞因子，在某些情况下也能被生长因子和GPCR兴奋剂激活(6,7)。二聚体SAPK/JNK被活化后能转位到细胞核中，在这里它能通过它对c-Jun, ATF-2和其他转录因子的效应而调节转录(8)。为了以持续的方式有效地促进生长和细胞分化，生长因子和有丝分裂原必须上调翻译(9,10)。生长因子和有丝分裂原能诱导 p70 S6 kinase的活化，，该激酶又能磷酸化S6核糖体蛋白。S6的磷酸化与翻译增加有关，特别是5+非翻译区域内带帽结构的mRNAs(10)。mRNAs (5'TOP)编码的蛋白涉及细胞周期进程和翻译机制部分的蛋白，如核糖体蛋白和延长因子(10,11)。热休克蛋白(HSP) 27是小HSPs中的一种，它能调节转录和后转录水平(12)。应答应激反应后，HSP27的表达水平提高了几倍，从而使细胞对抗不利的环境改变。HSP27丝氨酸（15, 78，82位点）被MAPKAP kinase 2磷酸化，这是p38 MAP激酶通路活化的结果(13,14)。

1. Heim, M.H. (1999) J. Recept. Signal. Transduct. Res. 19, 75-120.
2. Durbin, J.E. et al. (1996) Cell 84, 443-450.
3. Meraz, M.A. et al. (1996) Cell 84, 431-442.
4. Ihle, J.N. et al. (1994) Trends Biochem. Sci. 19, 222-227.
5. Frank, D.A. (1999) Mol. Med. 5, 432-456.
6. Davis, R.J. (1999) Biochem. Soc. Symp. 64, 1-12.
7. Kyriakis, J.M. and Avruch, J. (2001) Phisiol. Rev. 81, 807-869.
8. Whitmarsh, A.J. and Davis, R.J. (1998) Trends Biochem. Sci. 23, 481-485.
9. Dufner, A. and Thomas, G. (1999) Exp. Cell. Res. 253, 100-109.
10. Peterson, R.T. and Schreiber, S.L. (1998) Curr. Biol. 8, R248-R250.
11. Jefferies, H.B. et al. (1997) EMBO J. 16, 3693-3704.
12. Arrigo, A.P. and Landry, J. (1994) The Biology of Heat Shock Proteins and Molecular Chaperones. Cold Spring Harbor Laboratory Press, NY 335-373.
13. Landry, J. et al. (1992) J. Biol. Chem. 267, 794-803.
14. Rouse, J. et al. (1994) Cell 78, 1027-1037.

原厂资料：

Specificity / Sensitivity

Each phospho-antibody in this cocktail recognizes endogenous levels of only the phosphorylated form of its specific target. The Pin1 antibody detects endogenous levels of its target protein and is provided to control for protein loading.

Source / Purification

Antibodies are produced by immunizing animals with synthetic peptides. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.

Description

The PathScan® Multiplex Western Cocktail III offers a unique method to assay the activation of multiple pathways on one membrane without stripping and reprobing. This method saves the user valuable time, while increasing accuracy and minimizing reagent waste. The system allows the user to simultaneously detect levels of phospho-Stat1, phospho-SAPK/JNK, phospho-S6 ribosomal protein and phospho-HSP27. The cocktail also includes Pin1 antibody to control protein loading.

Background

Stat1, while activated in response to a large number of ligands, appears to be essential for responsiveness to IFN-alpha and IFN-gamma (1-3). 
 Phosphorylation of Stat1 at Tyr701 induces Stat1 dimerization, nuclear translocation and DNA binding (4). Stat1 has been found to be inappropriately activated in many tumors (5). 
 The stress-activated protein kinase/Jun-terminal kinase SAPK/JNK is potently and preferentially activated by a variety of environmental stresses, including UV and gamma radiation, ceramides, inflammatory cytokines and, in some instances, by growth factors and GPCR agonists (6,7). SAPK/JNK, when active as a dimer, can translocate to the nucleus where it regulates transcription through its effects on c-Jun, ATF-2 and other transcription factors (8). 
 To effectively promote growth and cell division in a sustained manner, growth factors and mitogens must upregulate translation (9,10). Growth factors and mitogens induce the activation of p70 S6 kinase, which in turn phosphorylate the S6 ribosomal protein. Phosphorylation of S6 correlates with an increase in translation, particularly of mRNAs with an oligopyrimidine tract in their 5' untranslated regions (10). 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 (10,11). 
 Heat shock protein (HSP) 27 is one of the small HSPs, regulated at both the transcriptional and posttranslational levels (12). In response to stress, the expression level of HSP27 increases several-fold to confer cellular resistance to the adverse environmental change. HSP27 is also phosphorylated at serines 15, 78 and 82 by MAPKAP kinase 2 as a result of p38 MAP kinase pathway activation (13,14).

1. Heim, M.H. (1999) J. Recept. Signal. Transduct. Res. 19, 75-120.
2. Durbin, J.E. et al. (1996) Cell 84, 443-450.
3. Meraz, M.A. et al. (1996) Cell 84, 431-442.
4. Ihle, J.N. et al. (1994) Trends Biochem. Sci. 19, 222-227.
5. Frank, D.A. (1999) Mol. Med. 5, 432-456.
6. Davis, R.J. (1999) Biochem. Soc. Symp. 64, 1-12.
7. Kyriakis, J.M. and Avruch, J. (2001) Phisiol. Rev. 81, 807-869.
8. Whitmarsh, A.J. and Davis, R.J. (1998) Trends Biochem. Sci. 23, 481-485.
9. Dufner, A. and Thomas, G. (1999) Exp. Cell. Res. 253, 100-109.
10. Peterson, R.T. and Schreiber, S.L. (1998) Curr. Biol. 8, R248-R250.
11. Jefferies, H.B. et al. (1997) EMBO J. 16, 3693-3704.
12. Arrigo, A.P. and Landry, J. (1994) The Biology of Heat Shock Proteins and Molecular Chaperones. Cold Spring Harbor Laboratory Press, NY 335-373.
13. Landry, J. et al. (1992) J. Biol. Chem. 267, 794-803.
14. Rouse, J. et al. (1994) Cell 78, 1027-1037.

注意事项：

Storage: Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM
NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not
aliquot the antibody