The Caspase-Glo® 9 Assay is a homogeneous luminescent assay that measures caspase-9 activity. The assay provides a proluminogenic caspase-9 substrate in a buffer system optimized for caspase activity, luciferase activity and cell lysis. The addition of a single Caspase-Glo® 9 Reagent in an "add-mix-read" format results in cell lysis, followed by caspase cleavage of the substrate and generation of a "glow-type" luminescent signal. The signal generated is proportional to the amount of caspase activity present. The Caspase-Glo® Reagent relies on the properties of a proprietary thermostable luciferase (Ultra-Glo™ Recombinant Luciferase), which generates the stable "glow-type" luminescent signal and improves performance across a wide range of assay conditions.
The system now includes a separate vial of a protease inhibitor, MG-132 Inhibitor, which may be used to reduce background, thus improving the performance of the Caspase-Glo® 9 Assay in cell-based applications.
Features and Benefits
- The homogeneous "add-mix-read" protocol makes the assay easy to automate; simply add an equal volume of reagent to sample volume.
- The low background luminescence results in excellent signal-to-noise ratios and superior sensitivity not achieved by other caspase formats, allowing assays to be performed in 96- or 384-well formats.
- No sample preparation or manipulation required, and no extended incubation times are necessary as with fluorescent-based assays. Maximum sensitivity is achieved in as little as 0.5–1 hour.
- The assay delivers excellent Z´ factors in cell and purified enzyme models.
- The Caspase-Glo® 9 Assay uses a luminogenic substrate containing the LEHD sequence, which has been shown to be selective for caspase-9. The assay includes an optional proteasome inhibitor (MG-132), which when added to the Caspase-Glo® 9 Reagent significantly reduces nonspecific background in cell-based assays.
- Multiplex with other cell-based assays from Promega.
Applications include caspase-9 activity determination, caspase-9 inhibition determination (IC50), and caspase-9 activation determination (EC50).
Using 100μl of Caspase-Glo® Reagent per assay in a 96-well format, Cat.# G8210 contains sufficient reagents for 25 assays; Cat.# G8211, 100 assays; Cat.# G8212, 1,000 assays, respectively. Using 25μl of Caspase-Glo® Reagent per assay in a 384-well format, Cat.# G8210 contains sufficient reagents for 100 assays; Cat.# G8211, 400 assays; Cat.# G8212, 4,000 assays, respectively.