PathScan? Phospho-Met (Tyr1349) Sandwich ELISA Kit

• 产品编号：CST-7896C      品牌：CST       原厂货号：7896C
• 产品分类：ELISA及其它免疫分析 > ELISA > ELISA检测试剂盒
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描述：

CST公司的PathScan® Phospho-Met (Tyr1349) Sandwich ELISA Kit用于检测内源Tyr1349磷酸化的Met蛋白的表达（如图１）。图2显示试剂盒的灵敏度。内部检测显示，本试剂盒能检测列举出的物种蛋白，但不排除用于其它物种同源蛋白检测的可能。PathScan® Phospho-Met (Tyr1349) Sandwich ELISA Kit利用固相双抗夹心酶联免疫检测法对内源性Tyr1349磷酸化的Met的表达进行检测。微孔中已经包被了Met兔抗体，与细胞裂解液共孵育后，磷酸化Met (Tyr1349)能被包被抗体捕获。多次洗板之后，加入生物素偶联磷酸化Met (Tyr1349) 兔检测抗体对被捕获的磷酸化Met蛋白的进行检测，然后加入HRP偶联链霉亲和素对检测抗体进行识别，最后加入HRP的底物TMB进行显色反应。按照相对光单位，发射光强度与Tyr1349磷酸化的Met蛋白的量成正比。Met作为肝细胞生长因子 (HGF, 亦作扩散因子) 的高亲和性酪氨酸激酶受体是一个二硫键连接的异二聚体，含有一个45 kDa的 α-亚基和一个145 kDa的 β-亚基 (1,2)。α-亚基和β-亚基的氨基端形成胞外区，β-亚基的其余部分跨细胞膜并且含有一个具有激酶活性的胞内区。Met和HGF相互作用会导致多个酪氨酸位点的自磷酸化，继而招募下游信号分子Gab1, c-Cbl和 PI3 K激酶（3）。这一系列基础性事件对于Met激酶相关的所有生物学功能十分重要。胞内Tyr1003位点的磷酸化对于Met的泛素化和降解是必须的（4）。激酶结构域Tyr1234/1235的磷酸化对于Met的激酶活性十分关键。胞内 Tyr1349的磷酸化为Met提供了Gab1 的直接结合位点(5)。在肾、结肠和乳腺癌中发现有Met水平和/或激酶活性的改变，因此Met是肿瘤治疗和诊断的一个靶点（6,7）。

1. Cooper, C.S. et al. (1984) Nature 311, 29-33.
2. Bottaro, D.P. et al. (1991) Science 251, 802-4.
3. Bardelli, A. et al. (1997) Oncogene 15, 3103-11.
4. Taher, T.E. et al. (2002) J Immunol 169, 3793-800.
5. Schaeper, U. et al. (2000) J Cell Biol 149, 1419-32.
6. Eder, J.P. et al. (2009) Clin Cancer Res 15, 2207-14.
7. Sattler, M. and Salgia, R. (2009) Update Cancer Ther 3, 109-118.

原厂资料：

Specificity / Sensitivity

CST's PathScan® Phospho-Met (Tyr1349) Sandwich ELISA Kit detects endogeneous levels of Met when phosphorylated at Tyr1349 (see Figure 1). 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.

Description

The PathScan® Phospho-Met (Tyr1349) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Met when phosphorylated at Tyr1349. A Met rabbit antibody has been coated onto the microwells. After incubation with cell lysates, phospho-Met (Tyr1349) is captured by the coated antibody. Following extensive washing, a biotinylated phospho-Met (Tyr1349) rabbit detection antibody is added to detect the captured phospho-Met protein. HRP-linked streptavidin is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The absorbance at 450nm is proportional to the quantity of Met phosphorylated at Tyr1349.

Background

Met, a high affinity tyrosine kinase receptor for hepatocyte growth factor (HGF, also known as scatter factor) is a disulfide-linked heterodimer made of 45 kDa α- and 145 kDa β-subunits (1,2). The α-subunit and the amino-terminal region of the β-subunit form the extracellular domain. The remainder of the β-chain spans the plasma membrane and contains a cytoplasmic region with tyrosine kinase activity. Interaction of Met with HGF results in autophosphorylation at multiple tyrosines, which recruit several downstream signaling components, including Gab1, c-Cbl, and PI3 kinase (3). These fundamental events are important for all of the biological functions involving Met kinase activity. The addition of a phosphate at cytoplasmic Tyr1003 is essential for Met protein ubiquitination and degradation (4). Phosphorylation at Tyr1234/1235 in the Met kinase domain is critical for kinase activation. Phosphorylation at Tyr1349 in the Met cytoplasmic domain provides a direct binding site for Gab1 (5). Altered Met levels and/or tyrosine kinase activities are found in several types of tumors, including renal, colon, and breast. Thus, Met is an attractive cancer therapeutic and diagnostic target (6,7).

1. Cooper, C.S. et al. (1984) Nature 311, 29-33.
2. Bottaro, D.P. et al. (1991) Science 251, 802-4.
3. Bardelli, A. et al. (1997) Oncogene 15, 3103-11.
4. Taher, T.E. et al. (2002) J Immunol 169, 3793-800.
5. Schaeper, U. et al. (2000) J Cell Biol 149, 1419-32.
6. Eder, J.P. et al. (2009) Clin Cancer Res 15, 2207-14.
7. Sattler, M. and Salgia, R. (2009) Update Cancer Ther 3, 109-118.

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).