Caspase-8 IETD-R110 Fluorometric HTS Assay Kit

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Catalog No : 30012-1, 30012-2, 30012-3

Caspase-8 IETD-R110 Fluorometric HTS Assay Kit is specifically designed for HTS-based assays. The kit provides a homogenous assay system for fast and highly sensitive detection of caspase-8 activity by fluorescence in enzymatic reaction or mammalian cells. The fluorogenic substrate (Ac-IETD)₂-R110 contains two IETD tetrapeptides and is completely hydrolyzed by the enzyme 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 (λ_Ex/λ_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.

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.

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

Kit Components:

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

Features:

• HTS-compatible: Single-step homogenous assay specifically designed for HTS-based detection.
• Fast: Fast enzyme kinetics.
• Sensitive: The enzymatic reaction forms intensely green fluorescent rhodamine 110 (R110) product.
• MSDS (PDF): MSDS 30012