PathScan? Phospho-Chk2 (Thr68) Sandwich ELISA Kit

• 产品编号：CST-7037C      品牌：CST       原厂货号：7037C
• 产品分类：ELISA及其它免疫分析 > ELISA > ELISA检测试剂盒
• 应用分类：

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描述：

CST的PathScan® Phospho-cdc2 (Tyr15) Sandwich ELISA Kit能够检测内源性苏氨酸（68位）磷酸化的Chk2蛋白。本试剂盒能够检测通过内部测试的特定种属蛋白，但也许能够检测其他种属的同源蛋白。PathScan® Phospho-Chk2 (Thr68) Sandwich ELISA Kit是一种固相酶联免疫吸附试剂盒，能够检测内源性苏氨酸68位磷酸化的Chk2蛋白。微孔板上已包被有一种phospho-Chk2 (Thr68) rabbit antibody。细胞裂解液孵育之后，phospho-Chk2蛋白被包被抗体捕获。洗涤之后，加入Chk2 mouse detection antibody检测已捕获的Chk2蛋白。使用抗鼠IgG和HRP-连接的抗体#7076识别结合的检测抗体。加入HRP底物(TMB)进行显色。显色过程中吸光度的强度和苏氨酸68位磷酸化的Chk2蛋白的量成正比。该试剂盒中的抗体具有特异性的固定组成。Chk2的氨基末端结构域包含有一个七丝氨酸或者苏氨酸残基(Ser19, Thr26, Ser28, Ser33, Ser35, Ser50 and Thr68)组成的系列，每个系列紧跟着谷氨酸(SQ 或TQ模体)。这些地方是已知的ATM/ATR激酶磷酸化的首选作用位点(4,5)。电离辐射(IR)引起DNA损伤之后，UV照射处理或者羟基脲处理，这个区域的68位苏氨酸和其他位点被ATM/ATR磷酸化(5-7)。因此，SQ/TQ簇结构域似乎具有调节功能。第68位苏氨酸的磷酸化是后续激活步骤的先决条件，激活归因于激酶结构域激活回路中Chk2第383位和387位苏氨酸残基的自体磷酸化(8)。

1. Allen, J.B. et al. (1994) Genes Dev. 8, 2401-2415.
2. Weinert, T.A. et al. (1994) Genes Dev. 8, 652-665.
3. Murakami, H. and Okayama, H. (1995) Nature 374, 817-819.
4. Kastan, M.B. and Lim, D.S. (2000) Nat. Rev. Mol. Cell Biol. 1, 179-186.
5. Matsuoka, S. et al. (2000) Proc. Natl. Acad. Sci. USA 97, 10389-10394.
6. Melchionna, R. et al. (2000) Nat. Cell Biol. 2, 762-765.
7. Ahn, J.Y. et al. (2000) Cancer Res. 60, 5934-5936.
8. Lee, C.H. and Chung, J.H. (2001) J. Biol. Chem. 276, 30537-30541.

原厂资料：

Specificity / Sensitivity

CST's PathScan® Phospho-Chk2 (Thr68) Sandwich ELISA Kit #7037 detects endogenous levels of Chk2 protein when phosphorylated at Thr68. 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-Chk2 (Thr68) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Chk2 when phosphorylated at Thr68. A phospho-Chk2 (Thr68) rabbit antibody has been coated onto the microwells. After incubation with cell lysates, phospho-Chk2 protein is captured by the coated antibody. Following extensive washing, a Chk2 mouse detection antibody is added to detect the captured Chk2 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 the absorbance for the developed color is proportional to the quantity of Chk2 phosphorylated at Thr68. 
 Antibodies in kit are custom formulations specific to kit.

Background

Chk2 is the mammalian orthologue of the budding yeast Rad53 and fission yeast Cds1 checkpoint kinases (1-3). The amino-terminal domain of Chk2 contains a series of seven serine or threonine residues (Ser19, Thr26, Ser28, Ser33, Ser35, Ser50 and Thr68) each followed by glutamine (SQ or TQ motif). These are known to be preferred sites for phosphorylation by ATM/ATR kinases (4,5). After DNA damage by ionizing radiation (IR), UV irradiation or hydroxyurea treatment, Thr68 and other sites in this region become phosphorylated by ATM/ATR (5-7). The SQ/TQ cluster domain, therefore, seems to have a regulatory function. Phosphorylation at Thr68 is a prerequisite for the subsequent activation step, which is attributable to autophosphorylation of Chk2 on residues Thr383 and Thr387 in the activation loop of the kinase domain (8). 

1. Allen, J.B. et al. (1994) Genes Dev. 8, 2401-2415.
2. Weinert, T.A. et al. (1994) Genes Dev. 8, 652-665.
3. Murakami, H. and Okayama, H. (1995) Nature 374, 817-819.
4. Kastan, M.B. and Lim, D.S. (2000) Nat. Rev. Mol. Cell Biol. 1, 179-186.
5. Matsuoka, S. et al. (2000) Proc. Natl. Acad. Sci. USA 97, 10389-10394.
6. Melchionna, R. et al. (2000) Nat. Cell Biol. 2, 762-765.
7. Ahn, J.Y. et al. (2000) Cancer Res. 60, 5934-5936.
8. Lee, C.H. and Chung, J.H. (2001) J. Biol. Chem. 276, 30537-30541.

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).