PathScan? Phospho-MEK1 (Ser217/221) Sandwich ELISA Kit

• 产品编号：CST-7175C      品牌：CST       原厂货号：7175C
• 产品分类：ELISA及其它免疫分析 > ELISA > ELISA检测试剂盒
• 应用分类：

其它组分：

描述：

CST的 PathScan® Phospho-MEK1 (Ser217/221) Sandwich ELISA Kit能检测内源性磷酸化MEK1蛋白水平。如图1所示，使用此kit#7175可以检测到，经PMA处理过的NIH/3T3细胞内的p-MEK1被显著激活了。然而用PathScan® Total MEK1 Sandwich ELISA Kit #7165检测到的MEK1总蛋白水平未发生变化。此试剂盒也可以在人293细胞中检测磷酸化MEK1蛋白水平。经内部测试确认，本试剂盒可以检测到所列物种中的蛋白。但对其他物种中的同源蛋白可能也可以检测。CST的 PathScan® Phospho-MEK1 (Ser217/221) Sandwich ELISA Kit是通过固相夹心酶联免疫吸附法（ELISA）来检测内源性的Ser217/221位磷酸化的MEK1蛋白的水平。先将MEK1 Mouse mAb包被在微孔板上。与细胞裂解物孵育后，MEK1总蛋白（磷酸化和非磷酸化）便可被包被的抗体捕获。清洗彻底后，加入Phospho-MEK1/2 Antibody检测磷酸化MEK1蛋白的水平。再加入Anti-Rabbit IgG, HRP偶联抗体 #7074*作为二抗来识别之前绑定的检测抗体。最后加入HRP底物—— TMB，进行显色。产生的吸光值与磷酸化的MEK1的数量成正比。*试剂盒中抗体是专为本试剂盒特制的。MEK1、MEK2,也被称为MAPK或Erk激酶，它们是双特异性蛋白激酶，在丝裂原活化蛋白激酶级联反应中发挥作用，控制细胞生长和分化(1-3)。MEK1、MEK2的活化是通过Raf样分子对两个丝氨酸（217 /221位点）的磷酸化而实现的，这两个丝氨酸位点位于亚结构域VIII的活化环内。多种生长因子和细胞因子以及膜去极化和钙内流都能激活MEK1/2(1-4)。组成型活化的MEK1/2能够诱导NIH/3T3细胞的转化和PC-12细胞的分化(4)。MEK蛋白通过将p44、p42 MAPK激酶上位于亚结构域VIII的活化环内的丝氨酸和苏氨酸磷酸化，激活p44、 p42 MAPK激酶。

1. Crews, C.M. et al. (1992) Science 258, 478-480.
2. Alessi, D.R. et al. (1994) EMBO J. 13, 1610-1619.
3. Rosen, L.B. et al. (1994) Neuron 12, 1207-1221.
4. Cowley, S. et al. (1994) Cell 77, 841-852.

原厂资料：

Specificity / Sensitivity

CST's PathScan® Phospho-MEK1 (Ser217/221) Sandwich ELISA Kit detects endogenous levels of phospho-MEK1 protein. A significant induction of MEK1 phosphorylation in PMA-treated NIH/3T3 cells can be detected using this kit, #7175. However, the level of total MEK1 protein detected by PathScan® Total MEK1 Sandwich ELISA Kit #7165 remains unchanged (Figure 1). This kit can also be used to detect phosphorylated MEK1 protein in human 293 cells. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

Description

CST's PathScan® Phospho-MEK1 (Ser217/221) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Phospho-MEK1 (Ser217/221) protein. MEK1 Mouse mAb has been coated onto the microwells. After incubation with cell lysates, total MEK1 protein (phospho- and nonphospho-) is captured by the coated antibody. Following extensive washing, a Phospho-MEK1/2 Antibody is added to detect the captured phospho-MEK1 protein. HRP-linked anti-rabbit antibody #7074* is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of phospho-MEK1 protein. 
 * Antibodies in this kit are custom formulations specific to the the kit.

Background

MEK1 and MEK2, also called MAPK or Erk kinases, are dual-specificity protein kinases that function in a mitogen activated protein kinase cascade controlling cell growth and differentiation (1-3). Activation of MEK1 and MEK2 occurs through phosphorylation of two serine residues at positions 217 and 221, located in the activation loop of subdomain VIII, by Raf-like molecules. MEK1/2 is activated by a wide variety of growth factors and cytokines and also by membrane depolarization and calcium influx (1-4). Constitutively active forms of MEK1/2 are sufficient for the transformation of NIH/3T3 cells or the differentiation of PC-12 cells (4). MEK activates p44 and p42 MAP kinase by phosphorylating both threonine and tyrosine residues at sites located within the activation loop of kinase subdomain VIII.

1. Crews, C.M. et al. (1992) Science 258, 478-480.
2. Alessi, D.R. et al. (1994) EMBO J. 13, 1610-1619.
3. Rosen, L.B. et al. (1994) Neuron 12, 1207-1221.
4. Cowley, S. et al. (1994) Cell 77, 841-852.

注意事项：

 12 8-well modules -Each module is designed to break apart for 8 tests.
**Kit should be stored at 4°C with the exception of 10X Cell Lysis Buffer, which is stored at –20°C (packaged separately).