Microplate Assays for Caspase Activity

A distinctive feature of the early stages of apoptosis is the activation of caspase enzymes, which participate in the cleavage of protein substrates and in the subsequent disassembly of the cell. We offer a series of caspase assays that allow the simple detection of active caspases in living cells in real time or in cellular lysates or extracts.

Live-cell assays

CellEvent Caspase-3/7 Green Detection Reagent is optimized for apoptosis analysis with a simple no-wash protocol that helps preserve delicate apoptotic cells. Live cells in microplate wells can be monitored in real time for apoptotic activity by measuring fluorescence intensity.

Caspase-3/7 assays

Microplate assay kits for caspase-3/7 measurement provide a choice of substrates with different wavelength emissions. Kits are available using rhodamine 110 or AMC to report caspase activity. Substrates are also available as bulk reagents or predispensed into microplate wells

Dose–response curve with CellEvent Caspase-3/7 Green Detection Reagent.
Dose–response curve with CellEvent Caspase-3/7 Green Detection Reagent. The percent positive for active caspase3/7 and EC50 were determined for staurosporine-treated U2OS cells using the reagents in the CellEvent Caspase-3/7 Green Detection Reagent.
Selection guide for microplate caspase assays
  CellEvent Caspase-3/7 Green Detection Reagent EnzChek Caspase-3 Assay Kit #1, Z-DEVD-AMC Substrate EnzChek Caspase-3 Assay Kit #2, Z-DEVD-R110 Substrate
Reporter DEVD conjugated to a nucleic acid-binding dye Z-DEVD-AMC substrate Z-DEVD-R110 substrate
Ex/Em (nm) 502/530 342/441 496/520
Live cell Yes No No
Lysate No Yes Yes
Enzyme prep No Yes Yes
Components Enzyme substrate for live and fixed cells Includes reference standard and inhibitor Includes reference standard and inhibitor
For Use with (Equipment) Fluorescence Microscope, High Content Instrument, Microplate Readers Microplate Readers Microplate Readers
Protocol outline
  1. Add diluted CellEvent Caspase-3/7 Green Detection Reagent to cells
  2. Incubate for 30 minutes
  3. Measure fluorescence
  1. Lyse cells, isolate supernatant.
  2. Prepare DEVD substrate and apply to wells.
  3. Add samples to wells.
  4. Incubate samples.
  5. Measure fluorescence.
  6. Determine caspase activity using standard curve.
  1. Lyse cells, isolate supernatant
  2. Prepare DEVD substrate and apply to wells.
  3. Add samples to wells.
  4. Incubate samples.
  5. Measure fluorescence.
  6. Determine caspase activity using standard curve.
Format 25 µL 100 µL 500 assays using 100 µL reaction volume 500 assays using 100 µL reaction volume
Cat. No. C10723 C10423 E13183 E13184

Resources

Molecular Probes Handbook

BioProbes Journal Articles

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