PathScan? Apoptosis and Proliferation Multiplex IF Kit

产品编号：CST-7851S 品牌：CST 原厂货号：7851S
产品分类：生化和细胞分析 > 细胞分析试剂盒 > 细胞凋亡试剂盒
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–20°C.

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其它组分：

PathScan® Multiplex IF Kit Detection Cocktail III【包装：100 µl，，运保温度：–20°C.】

PathScan® Apoptosis and Proliferation Multiplex IF Kit Primary Antibody Cocktail【包装：100 µl，，运保温度：–20°C.】

描述：

α-Tubulin antibody 能够检测内源α-tubulin总蛋白。Phospho-Histone H3 (Ser10) antibody仅能够检测内源性的Ser10磷酸化的histone H3蛋白，该抗体不会与其它磷酸化或乙酰化组蛋白产生交叉反应。Cleaved-PARP (Asp214) 能够检测内源性经caspase酶切的人源PARP1蛋白大片段(89 kDa)，该抗体不与全长PARP1或其它的PARP亚型发生交叉反应。试剂盒里的单克隆抗体是分别采用合成的人源组蛋白H3的Ser10周围残基相应的磷酸化肽段、人源PARP蛋白Asp213 周围残基相对应的磷酸化肽段或大脑提取物中纯化的全长鸡α-tubulin免疫动物而制备的。PathScan® Apoptosis and Proliferation Multiplex IF kit提供了一种新的方法，借助于人工免疫荧光显微镜或自动成像和激光扫描高通量平台，可以同时监控有丝分裂指数和细胞程序性死亡。此试剂盒包含三个高品质一抗靶向α-微管蛋白，磷酸化组蛋白H3（Ser10）和cleaved PARP（Asp214），并利用Alexa Fluor®系列荧光染料进行检测。抗体和染料配对已经预先优化，每个试剂盒中的试剂足够用于100次检测（基于工作体积为100μl/测试）。细胞凋亡是一个被调控的生理过程，导致细胞死亡。引发剂caspases切割并激活下游的效应caspases，它们反过来切割细胞骨架和核蛋白质，如PARP、α-胞衬蛋白、DFF和lamin A，并诱导细胞凋亡（1）。核聚（ADP-核糖）聚合酶PARP的裂解发生在Asp214和Gly215之间，这将PARP氨基末端的DNA结合域（24 kDa）从羧基端的催化结构域（89 kDa）分离开（2,3）。PARP有助于维持细胞活力，在许多细胞过程中起着关键作用，包括DNA复制、修复和重组。PARP的剪切促进细胞拆分，并作为细胞发生凋亡的一个标记（4）。可以通过研究组蛋白H3的磷酸化状态和有丝分裂过程中微管组装情况来测量细胞的增殖。组蛋白H3是四个不同的核心组蛋白之一，发现于核小体中。组蛋白H3 在Ser10、Ser28和Thr11处的磷酸化与有丝分裂和减数分裂时染色体的凝聚紧密相关（5-7）。 α-微管蛋白和β-微管蛋白组成的异源二聚体形成的球状微管蛋白亚基，是所有真核细胞共有的。被称为微管的微管蛋白聚合物是基础的胞浆纤维，对于介导细胞运动包括减数分裂/有丝分裂染色体排列，胞质膜泡运输，以及神经细胞轴突迁移等都非常重要（8）。

原厂资料：

Specificity / Sensitivity

α-Tubulin antibody detects endogenous levels of total α-tubulin protein. Phospho-Histone H3 (Ser10) antibody detects endogenous levels of histone H3 only when phosphorylated at Ser10. This antibody does not cross-react with other phosphorylated or acetylated histones. Cleaved-PARP (Asp214) detects endogenous levels of the large fragment (89 kDa) of human PARP1 protein produced by caspase cleavage. This antibody does not recognize full length PARP1 or other PARP isoforms.

Source / Purification

Monoclonal antibodies were produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser10 of human histone H3, a synthetic peptide corresponding to residues surrounding Asp213 of human PARP, or with full-length chicken α-tubulin purified from brain extracts.

Description

The PathScan® Apoptosis and Proliferation Multiplex IF kit offers a novel method to simultaneously monitor mitotic index and programmed cell death using manual immunofluorescence microscopy, or automated imaging and laser scanning high content platforms. This kit contains a cocktail of three high quality primary antibodies targeted against α-tubulin, phospho-histone H3 (Ser10), and cleaved PARP (Asp214), as well as a detection cocktail utilizing the Alexa Fluor® series of fluorescent dyes. Antibody and dye pairings have been pre-optimized, and each kit contains enough reagents for 100 assays (based on a working volume of 100 μL/test).

Background

Apoptosis is a regulated physiological process leading to cell death. Initiator caspases cleave and activate downstream effector caspases that in turn cleave cytoskeletal and nuclear proteins like PARP, α-fodrin, DFF, and lamin A, which induce apoptosis (1). Cleavage of the nuclear poly (ADP-ribose) polymerase PARP occurs between Asp214 and Gly215, which separates the PARP amino-terminal DNA binding domain (24 kDa) from the carboxy-terminal catalytic domain (89 kDa) (2,3). PARP helps to maintain cell viability by playing key roles in many cellular processes, including DNA replication, repair, and recombination. Cleavage of PARP facilitates cellular disassembly and serves as a marker of cells undergoing apoptosis (4). Cell proliferation can be measured by studying the phosphorylation state of histone H3 and microtubule assembly during mitosis. Histone H3 is one of four distinct core histone proteins found in the nucleosome. Phosphorylation of histone H3 at Ser10, Ser28, and Thr11 is tightly correlated with chromosome condensation during both mitosis and meiosis (5-7). Heterodimers composed of α-tubulin and β-tubulin form globular tubulin subunits common to all eukaryotic cells. Tubulin polymers known as microtubules are fundamental cytosolic fibers important in mediating cellular movement, including meiotic/mitotic chromosome alignment, cytoplasmic membrane vesicle transport, and nerve-cell axon migration (8).

注意事项：

Storage: Supplied in 10 mM sodium HEPES (pH 7.5), 150
mM NaCl, 100 μg/ml BSA, 50% glycerol and less than 0.02%
sodium azide. Store at –20°C. Do not aliquot either cocktail.