PathScan? Phospho-Stat5 (Tyr694) Sandwich ELISA Kit

• 产品编号：CST-7113C      品牌：CST       原厂货号：7113C
• 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的PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit #7113检测内源水平在Tyr694位点磷酸化Stat5a和Tyr699位点磷酸化的Stat5b蛋白。如图1所示，在HeLa细胞中，用Phospho-Stat5 (Tyr694) Sandwich ELISA Kit #7113检测经EGF处理诱导磷酸化Stat5蛋白。通过Western免疫印迹实验表明总Stat5蛋白并未受到影响。此试剂盒能够检测已经经内部严格测试的物种的蛋白，但是也不排除检测到其它物种中同源性的蛋白。CST的PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit是固相夹心酶联免疫吸收剂检测试剂盒(ELISA)，它能够检测在Tyr694位点磷酸化Stat5a和Tyr699位点磷酸化的Stat5b蛋白。Phospho-Stat5鼠抗包裹到微孔的外表面。然后与细胞裂解液孵育，phospho-Stat5a (Tyr694) 和 phospho-Stat5b (Tyr699)蛋白能够被抗体捕获。经过严格的冲洗后，Stat5兔检测抗体加入用来检测捕捉到的phospho-Stat5a和phospho-Stat5b蛋白。抗兔IgG，HRP连接抗体#7074用于识别检测抗体的结合。HRP的底物TMB加入进行颜色反应。对反应的颜色，光密度的数值按比例换算成在Tyr694位点磷酸化Stat5a和Tyr699位点磷酸化的Stat5b蛋白的量。*试剂盒中的抗体是经过客户证实试剂盒特异的。Stat5的激活是通过一系列配体的结合而完成，包括 IL-2、GM-CSF、生长激素和泌乳刺激素。磷酸化Tyr694位点对 Stat5的活性是必须的(1,2)。此位点的磷酸化是在红细胞生成素的作用下由Src介导而完成(3)。 Stat5在某些白血病类型的细胞中有组成性的活性(4)。用IL-3处理的内皮细胞中有磷酸化的Stat5，这就表明它参与了血管再生和细胞的运动(5)。在不同细胞类型中 Stat5a 和 Stat5b 是被独立调控和激活的。例如,干扰素处理后在U-937细胞中Stat5a的活性较强但在HeLa细胞中Stat5b的活性较强(6)。

1. Gouilleux, F. et al. (1994) EMBO J 13, 4361-9.
2. Wakao, H. et al. (1994) EMBO J 13, 2182-91.
3. Okutani, Y. et al. (2001) Oncogene 20, 6643-50.
4. Demoulin, J.B. et al. (1999) J Biol Chem 274, 25855-61.
5. Dentelli, P. et al. (1999) J Immunol 163, 2151-9.
6. Meinke, A. et al. (1996) Mol Cell Biol 16, 6937-44.

原厂资料：

Specificity / Sensitivity

CST’s PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit #7113 detects endogenous levels of phospho-Stat5a when phosphorylated at Tyr694 and Stat5b at Tyr699. As shown in Figure 1, a significant induction of Stat5 phosphorylation can be detected in EGF-treated HeLa cells using the Phospho-Stat5 (Tyr694) Sandwich ELISA Kit #7113. The level of total Stat5 detected by western analysis remains unchanged. 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-Stat5 (Tyr694) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Stat5a when phosphorylated at Tyr694 and Stat5b at Tyr699. A Phospho-Stat5 Mouse Antibody has been coated onto the microwells. After incubation with cell lysates, phospho-Stat5a (Tyr694) and phospho-Stat5b (Tyr699) proteins are captured by the coated antibody. Following extensive washing, a Stat5 Rabbit Detection Antibody is added to detect the captured phospho-Stat5a and phospho-Stat5b protein. Anti-rabbit IgG, HRP-linked antibody #7074 is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of absorbance for this developed color is proportional to the quantity of Stat5a phosphorylated at Tyr694 and Stat5b phosphorylated at Tyr699. 
 Antibodies in kit are custom formulations specific to kit.

Background

Stat5 is activated in response to a wide variety of ligands including IL-2, GM-CSF, growth hormone and prolactin. Phosphorylation at Tyr694 is obligatory for Stat5 activation (1,2). This phosphorylation is mediated by Src upon erythropoietin stimulation (3). Stat5 is constitutively active in some leukemic cell types (4). Phosphorylated Stat5 is found in some endothelial cells treated with IL-3, which suggests its involvement in angiogenesis and cell motility (5). Stat5a and Stat5b are independently regulated and activated in various cell types. For instance, interferon treatment predominantly activates Stat5a in U-937 cells and Stat5b in HeLa cells (6).

1. Gouilleux, F. et al. (1994) EMBO J 13, 4361-9.
2. Wakao, H. et al. (1994) EMBO J 13, 2182-91.
3. Okutani, Y. et al. (2001) Oncogene 20, 6643-50.
4. Demoulin, J.B. et al. (1999) J Biol Chem 274, 25855-61.
5. Dentelli, P. et al. (1999) J Immunol 163, 2151-9.
6. Meinke, A. et al. (1996) Mol Cell Biol 16, 6937-44.

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).