PathScan? Phospho-ALK (Tyr1604) Sandwich ELISA Kit

• 产品编号：CST-7324C      品牌：CST       原厂货号：7324C
• 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公司的PathScan®磷酸化ALK(Tyr1604)双抗夹心法化学发光检测试剂盒#7324主要用于检测Tyr1604磷酸化的ALK蛋白和磷酸化融合蛋白NPM-ALK的表达。如图1示，ALK和NPM-ALK融合蛋白在Karpas299细胞中被组成性地磷酸化，因此在该细胞系中能检测到磷酸化ALK (Tyr1604)和磷酸化NPM-ALK融合蛋白的高水平表达。这种高水平的表达在Karpas299细胞裂解液不加入磷酸酶抑制剂* 时被消除。而用PathScan®总ALK双抗夹心法酶联免疫检测试剂盒#7322对总ALK (磷酸化和非磷酸化) 的检测结果没有改变。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。*，磷酸酶抑制剂，包括焦磷酸钠、β-甘油磷酸二钠和Na3VO4。CST公司的PathScan®磷酸化-ALK(Tyr1604)双抗夹心法酶联免疫检测试剂盒利用固相双抗夹心酶联免疫检测法（ELISA）对Tyr1604磷酸化的ALK蛋白和融合蛋白NPM-ALK的表达进行检测。微孔中已经包被了抗磷酸化ALK(Tyr1604)的抗体，与细胞裂解液共孵育后，磷酸化的ALK和磷酸化的NPM-ALK融合蛋白能够被包被抗体捕获。多次洗板之后，加入另一种抗ALK的小鼠源抗体对磷酸化的ALK和磷酸化的NPM-ALK融合蛋白进行检测，然后加入HRP标记的抗小鼠的IgG#7076对检测抗体进行识别，最后加入HRP的底物TMB检测信号。按照相对光单位，发射光强度与Tyr1604磷酸化的ALK和NPM-ALK融合蛋白的量成正比。试剂盒中的抗体为试剂盒特异定制配方。间变性淋巴瘤激酶（ALK）是多效生长因子（PTN）的酪氨酸激酶受体，它是大脑胚胎发育过程中的生长因子(1-3)。 在表达ALK的细胞中，PTN诱导ALK及其下游效应因子IRS-1, Shc, PLCγ, 和PI3K激酶发生磷酸化(1)。ALK最初是从易位突变产生的NPM-ALK融合蛋白而被发现的(4)。NPM-ALK融合蛋白是一个组成性活化，有原癌基因活性的间变性淋巴瘤相关酪氨酸激酶(4)。 由NPM-ALK介导的PLCγ的活化可能对有丝分裂活性起到关键作用，并且可能对间变性淋巴瘤的发病过程很重要(5)。另一个完全不同的ALK原癌融合蛋白在非小细胞肺癌细胞株中被报道，该蛋白融合了ALK与EML4，其相应融合转录产物在一些肺腺癌中也有表达。在EML4-ALK融合蛋白中，微管相关蛋白EML4的短氨基末端区域和ALK的激酶活性区域融合在一起(6,7)。

1. Stoica, G.E. et al. (2001) J Biol Chem 276, 16772-9.
2. Iwahara, T. et al. (1997) Oncogene 14, 439-49.
3. Morris, S.W. et al. (1997) Oncogene 14, 2175-88.
4. Morris, S.W. et al. (1994) Science 263, 1281-4.
5. Bai, R.Y. et al. (1998) Mol Cell Biol 18, 6951-61.
6. Rikova, K. et al. (2007) Cell 131, 1190-203.
7. Takeuchi, K. et al. (2008) Clin Cancer Res 14, 6618-24.
8. Soda, M. et al. (2007) Nature 448, 561-6.

原厂资料：

Specificity / Sensitivity

CST's PathScan® Phospho-ALK (Tyr1604) Sandwich ELISA Kit #7324 detects endogenous levels of phospho-ALK (Tyr1604) protein or phospho-NPM-ALK fusion protein. As shown in Figure 1, a high level of phosphorylated ALK (Tyr1604) protein or phospho-NPM-ALK fusion protein is detected in Karpas299 cells where ALK or NPM-ALK is constitutively phosphorylated. These high levels are abolished in Karpas299 cells lysed without addition of phosphatase inhibitors* to the lysis buffer. The levels of total ALK protein (phospho and nonphospho) detected by PathScan® Total ALK Sandwich ELISA Kit #7322 remain unchanged. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species. 
 * Phosphatase inhibitors includes sodium pyrophosphate, β-glycerophosphate and Na3VO4.

Description

CST's PathScan® Phospho-ALK (Tyr1604) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of phospho-ALK (Tyr1604) or phospho-NPM-ALK fusion protein. A Phospho-ALK (Tyr1604) Antibody has been coated onto the microwells. After incubation with cell lysates, only phospho-ALK or phospho-NPM-ALK proteins are captured by the coated antibody. Following extensive washing, an ALK Mouse mAb is added to detect the captured phospho-ALK or phospho-NPM-ALK fusion protein. Anti-mouse IgG, HRP-linked Antibody #7076 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 phospho-ALK (Tyr1604) or phospho-NPM-ALK proteins. 
 Antibodies in kit are custom formulations specific to kit.

Background

Anaplastic lymphoma kinase (ALK) is a tyrosine kinase receptor for pleiotrophin (PTN), a growth factor involved in embryonic brain development (1-3). In ALK-expressing cells, PTN induces phosphorylation of both ALK and the downstream effectors IRS-1, Shc, PLCγ, and PI3 kinase (1). ALK was originally discovered as an NPM (nucleophosmin)-ALK fusion protein produced by a translocation (4). The NPM-ALK fusion protein is a constitutively active, oncogenic tyrosine kinase associated with anaplastic lymphoma (4). Activation of PLCγ by NPM-ALK has been suggested to be a crucial step for its mitogenic activity and may be important in the pathogenesis of anaplastic lymphomas (5). A distinct ALK oncogenic fusion protein involving ALK and EML4 has been described from a non-small cell lung cancer cell line, with corresponding fusion transcripts present in some cases of lung adenocarcinoma. The short, amino-terminal region of the microtubule-associated protein EML4 is fused to the kinase domain of ALK (6,7).

1. Stoica, G.E. et al. (2001) J Biol Chem 276, 16772-9.
2. Iwahara, T. et al. (1997) Oncogene 14, 439-49.
3. Morris, S.W. et al. (1997) Oncogene 14, 2175-88.
4. Morris, S.W. et al. (1994) Science 263, 1281-4.
5. Bai, R.Y. et al. (1998) Mol Cell Biol 18, 6951-61.
6. Rikova, K. et al. (2007) Cell 131, 1190-203.
7. Takeuchi, K. et al. (2008) Clin Cancer Res 14, 6618-24.
8. Soda, M. et al. (2007) Nature 448, 561-6.

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).