PathScan? Hydroxy-HIF-1α (Pro564) Sandwich ELISA Kit

• 产品编号：CST-13201C      品牌：CST       原厂货号：13201C
• 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：

如图1所示， PathScan^® Hydroxy-HIF-1α (Pro564) Sandwich ELISA试剂盒可以识别内源性的hydroxy-HIF-1α (Pro564)蛋白。试剂盒的敏感性如图2所示。内部测试表明，该试剂盒能检测来自指定物种的蛋白，但也可能会检测到来自其他物种的同源蛋白。PathScan®Hydroxy-HIF-1α (Pro564) Sandwich ELISA试剂盒是一个固相三明治酶联免疫反应实验(ELISA)能够检测内源性的hydroxy-HIF-1α (Pro564)蛋白。将HIF-1α抗体包被到微孔中。与细胞裂解液孵育后，HIF-1α蛋白会被抗体捕获。经过多次冲洗，再加入Hydroxy-HIF-1α (Pro564) 检测抗体以检测捕获到的hydroxy-HIF-1α (Pro564) 蛋白。Anti-rabbit IgG, HRP偶联的抗体随后用于检测结合上去的抗体。加入HRP底物，TMB，产生颜色。检测到的颜色的强度与hydroxy-HIF-1α (Pro564)蛋白量成正比。试剂盒中的抗体是专门为本试剂盒的使用所研发的。

缺氧诱导因子1(HIF1)是一个二聚体转录因子在细胞应激缺氧环境时会发挥作用(1)。HIF1复合物包括两个亚基，HIF-1α和HIF-1β，都属于PAS(Per, ARNT, Sim)碱性螺旋-环-螺旋家族(2)。HIF1调控多种应激缺氧环境的基因，包括调控血管生成，红细胞生成，细胞周期，细胞代谢和细胞凋亡过程的基因。广泛表达的HIF-1α通常是由泛素/蛋白酶体途径迅速在含氧量正常的细胞中降解的。在常氧条件下，HIF-1α被脯氨酸羟基化促进了其被绑定到von Hippel Lindau蛋白(VLH)E3连接酶复合物引起构象变化，随后被泛素化和蛋白酶体降解(3,4)。低氧条件和化学羟化酶抑制剂(如去铁胺和钴)抑制HIF-1α的降解，并增加其蛋白稳定性。此外，HIF-1α可通过各种细胞因子经过PI3K-AKT-mTOR通路诱导进入氧无关的信号通路(5-7)。

HIF-1β由于能够以伴侣方式与芳香烃受体(AHR)结合形成异二聚体复合物，也被称为芳香烃受体核转位(ARNT) (8)。HIF-1β与AhR能够在外源性化学物质代谢中发挥重要作用(8)。除此以外染色体转位导致的TEL-ARNT融合蛋白和急性髓细胞白血病相关(9)。研究也发现ARNT/HIF-1β的表达水平在2型糖尿病人的胰岛内明显表达下降，提示HIF-1β在胰岛β细胞功能过程中发挥作用(10)。

1. Sharp, F.R. and Bernaudin, M. (2004) Nat Rev Neurosci 5, 437-48.
2. Wang, G.L. et al. (1995) Proc Natl Acad Sci U S A 92, 5510-4.
3. Jaakkola, P. et al. (2001) Science 292, 468-72.
4. Maxwell, P.H. et al. (1999) Nature 399, 271-5.
5. Fukuda, R. et al. (2002) J Biol Chem 277, 38205-11.
6. Jiang, B.H. et al. (2001) Cell Growth Differ 12, 363-9.
7. Laughner, E. et al. (2001) Mol Cell Biol 21, 3995-4004.
8. Walisser, J.A. et al. (2004) Proc Natl Acad Sci U S A 101, 16677-82.
9. Salomon-Nguyen, F. et al. (2000) Proc Natl Acad Sci U S A 97, 6757-62.
10. Gunton, J.E. et al. (2005) Cell 122, 337-49.

原厂资料：

Specificity / Sensitivity

PathScan^® Hydroxy-HIF-1α (Pro564) Sandwich ELISA Kit recognizes endogenous levels of hydroxy-HIF-1α (Pro564) protein, as shown in 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^® Hydroxy-HIF-1α (Pro564) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of hydroxy-HIF-1α (Pro564) protein. A HIF-1α antibody has been coated onto the microwells. After incubation with cell lysates, HIF-1α protein is captured by the coated antibody. Following extensive washing, a Hydroxy-HIF-1α (Pro564) Detection Antibody is added to detect the captured hydroxy-HIF-1α (Pro564) protein. Anti-rabbit IgG, HRP-linked antibody 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 hydroxy-HIF-1α (Pro564) protein.Antibodies in kit are custom formulations specific to kit.

Background

Hypoxia-inducible factor 1 (HIF1) is a heterodimeric transcription factor that plays a critical role in the cellular response to hypoxia (1). The HIF1 complex consists of two subunits, HIF-1α and HIF-1β, which are basic helix-loop-helix proteins of the PAS (Per, ARNT, Sim) family (2). HIF1 regulates the transcription of a broad range of genes that facilitate responses to the hypoxic environment, including genes regulating angiogenesis, erythropoiesis, cell cycle, metabolism, and apoptosis. The widely expressed HIF-1α is typically degraded rapidly in normoxic cells by the ubiquitin/proteasomal pathway. Under normoxic conditions, HIF-1α is proline hydroxylated leading to a conformational change that promotes binding to the von Hippel Lindau protein (VLH) E3 ligase complex; ubiquitination and proteasomal degradation follows (3,4). Both hypoxic conditions and chemical hydroxylase inhibitors (such as desferrioxamine and cobalt) inhibit HIF-1α degradation and lead to its stabilization. In addition, HIF-1α can be induced in an oxygen-independent manner by various cytokines through the PI3K-AKT-mTOR pathway (5-7).

HIF-1β is also known as AhR nuclear translocator (ARNT) due to its ability to partner with the aryl hydrocarbon receptor (AhR) to form a heterodimeric transcription factor complex (8). Together with AhR, HIF-1β plays an important role in xenobiotics metabolism (8). In addition, a chromosomal translocation leading to a TEL-ARNT fusion protein is associated with acute myeloblastic leukemia (9). Studies also found that ARNT/HIF-1β expression levels decrease significantly in pancreatic islets from patients with type 2 diabetes, suggesting that HIF-1β plays an important role in pancreatic β-cell function (10).

1. Sharp, F.R. and Bernaudin, M. (2004) Nat Rev Neurosci 5, 437-48.
2. Wang, G.L. et al. (1995) Proc Natl Acad Sci U S A 92, 5510-4.
3. Jaakkola, P. et al. (2001) Science 292, 468-72.
4. Maxwell, P.H. et al. (1999) Nature 399, 271-5.
5. Fukuda, R. et al. (2002) J Biol Chem 277, 38205-11.
6. Jiang, B.H. et al. (2001) Cell Growth Differ 12, 363-9.
7. Laughner, E. et al. (2001) Mol Cell Biol 21, 3995-4004.
8. Walisser, J.A. et al. (2004) Proc Natl Acad Sci U S A 101, 16677-82.
9. Salomon-Nguyen, F. et al. (2000) Proc Natl Acad Sci U S A 97, 6757-62.
10. Gunton, J.E. et al. (2005) Cell 122, 337-49.

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).