Caspase-8 IETD-R110 荧光HTS检测试剂盒

Caspase-8 IETD-R110 荧光HTS分析试剂盒用于HTS为基础的分析。试剂盒提供一种快速高灵敏的方法荧光检测酶促反应或哺乳细胞的caspase-8活性。底物(Ac-IETD)2-R110 包含2个 IETD四肽，可以被两个连续步骤中的酶完全水解。第一个IETD肽段断裂产生单肽Ac-IETD-R110中间产物，吸收和发射波长和R110相似，但是荧光是后者的10%。第二个肽段水解释放出R110，形成大量荧光。试剂盒包含caspase-8抑制剂Ac-IETD-CHO,，可作为阴性对照，R110可用于做标准曲线，可以对caspase-8活性进行定量。

Caspase-3 DEVD-R110 Fluorometric HTS Assay Kit is a homogenous assay system for fast and highly sensitive detection of caspase-3 activity in mammalian cells. The kit is specifically designed for high throughput screening (HTS)-based assays.

Features

• Caspase-8 activity measured with substrate (Ac-IETD)2-R110
• Designed for high throughout assays
• Fast enzyme kinetics
• Enzymatic reaction forms green fluorescent R110 product
• Endpoint assay requires cell lysis

Kit Components

• Cell lysis/assay buffer
• Enzyme substrate (Ac-IETD)2-R110
• Enzyme inhibitor Ac-IETD-CHO
• R110

About this kit

Caspase-8 is the most upstream caspase in the CD95/Fas apoptotic pathway and is activated by the signaling pathways for CD95/Fas and TNF. The fluorogenic and chromogenic substrate (Ac-IETD)2-R110 contains two IETD tetrapeptides and is completely hydrolyzed by the caspase-8 in two successive steps. Cleavage of the first IETD peptide results in the monopeptide Ac-IETD-R110 intermediate, which has absorption and emission wavelengths similar to those of R110 (rhodamine 110) (Ex/Em= 496/520 nm) but has only about 10% of the fluorescence of the latter. Hydrolysis of the second IETD peptide releases the dye R110, leading to a substantial fluorescence increase.

This assay kit includes Ac-IETD-CHO, which is a caspase-8 inhibitor and can be used as a negative control. R110 is also provided in the kit for generating a standard curve, which can be used for quantifying caspase-8 activity.

Note: While caspase-8 preferentially cleaves the consensus sequence IETD compared to other substrate sequences, other caspases such as caspase-3 also can cleave IETD efficiently. Overlapping caspase substrate recognition limits the usefulness of caspase substrate peptides for distinguishing between different caspase activities in cell lysates.