These Fluorometric and Colorimetric Assay Kits provide a simple assay system for fast and highly sensitive detection of Caspase-3 or Caspase-8 activity either by fluorescence or absorbance in mammalian cells.
- Fast, sensitive enzyme kinetics
- Enzymatic reaction forms intensely yellow coloured and highly green fluorescent rhodamine 110 (R110) product
- Compatible with both fluorometric and colorimetric detection systems
- Single-step homogenous assay specifically designed for high-throughput screening (HTS)-based detection
Caspase-3 is an active cell-death protease involved in the execution phase of apoptosis, during which cells undergo morphological changes such as DNA fragmentation, chromatin condensation and apoptotic body formation. Caspase-3 DEVD-R110 kits provide a simple assay system for fast and highly sensitive detection of caspase-3 activity either by fluorescence or absorbance in mammalian cells. The fluorogenic and chromogenic substrate (Ac-DEVD)2-R110 contains two DEVD tetrapeptides and is completely hydrolyzed by the enzyme in two successive steps.
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. Caspase-8 kits provide a simple assay system for fast and sensitive detection of caspase-8 activity either by fluorescence or absorbance. The substrate (Ac-IETD)2-R110 contains two IETD tetrapeptides and is completely hydrolyzed by the enzyme in a two-step process.
Although fluorometric detection of the end products is preferred because of the superior sensitivity, detection by absorbance is also possible. In fact, the extinction coefficient of R110 is 10 times higher than that of p-nitroaniline (pNA), a dye commonly used in chromogenic substrates, making R110-based substrates significantly more sensitive than pNA-based substrates, even by colorimetric detection.
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.
Informace o dodávce: Kit includes cell lysis buffer, assay buffer, enzyme substrate (Ac-DEVD)2-R110 or (Ac-IETD)2-R110, enzyme inhibitor Ac-DEVD-CHO or Ac-IETD-CHO, R110 (for generating a standard curve for quantifying caspase-3 activity).