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 Detection Reagent is optimized for apoptosis analysis with a simple no-wash protocol that helps preserve delicate apoptotic cells, it is available in both green and red fluorescent forms. 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

 

CellEvent Caspase-3/7 Red vs CellEvent Caspase-3/7 Green Dose Response Curve with fluorescence microplate reader. A673 cells were seeded at 5,000 cells per well and treated with staurosporine at the indicated concentration for five hours at 37°C. Cells induced for apoptosis were stained with powder stock formulations of CellEvent Red or Green Reagent prepared in PBS and added directly to the cultures for 1 hour at 37°C prior to scanning fluorescence signal on the Varioskan LUX multimode microplate reader.

Selection guide for microplate caspase assays
 CellEvent Caspase-3/7 Green Detection ReagentCellEvent Caspase-3/7 Red Detection Reagent  EnzChek Caspase-3 Assay Kit #1, Z-DEVD-AMC SubstrateEnzChek Caspase-3 Assay Kit #2, Z-DEVD-R110 Substrate
ReporterDEVD conjugated to a nucleic acid-binding dyeDEVD conjugated to a nucleic acid-binding dyeZ-DEVD-AMC substrateZ-DEVD-R110 substrate
Ex/Em (nm)502/530590/610 342/441496/520
Live cellYesYesNoNo
LysateNo NoYesYes
Enzyme prepNo NoYesYes
ComponentsEnzyme substrate for live and fixed cellsEnzyme substrate for live and fixed cellsIncludes reference standard and inhibitorIncludes reference standard and inhibitor
For use with (Equipment)Fluorescence microscope, High-content instrument, Microplate readersFluorescence microscope, High-content instrument, Microplate readersMicroplate readersMicroplate readers
Protocol outline
  1. Add diluted CellEvent Caspase-3/7 Green Detection Reagent to cells
  2. Incubate for 30 minutes
  3. Measure fluorescence
  1. Add diluted CellEvent Caspase-3/7 Red 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
Format25 µL
(400X in DMSO)		100 µL
(400X in DMSO)		1 vial (good for 100 96-well assays)5 vials (good for 500 96-well assays)  500 assays using 100 µL reaction volume500 assays using 100 µL reaction volume
Cat. No.C10723C10423C10430C10431  E13183E13184

Expertly detect fluorescence with Thermo Scientific plate readers

product photo

High-sensitivity fluorescence detection for 6-1,536 samples can be quickly performed on the Varioskan ALF or Varioskan LUX Multimode Microplate Reader using Invitrogen reagents to enable optimal detection. Take advantage of automatic dynamic range selection to get optimal gain settings for each individual well and automation capabilities for even higher throughput.

Compare readers

Resources

For Research Use Only. Not for use in diagnostic procedures.