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. Life Technologies offers a series of caspase assays that allow the simple detection of active caspases in living cells in real time or as a snapshot.

Select caspase activity assays

Also see flow cytometry assays for caspase activity

Caspase Imaging Assays

  • Real-time no-wash assay for caspase activity
  • Multiparameter assays for live and dead cells
  • Antibodies for immunodetection of apoptosis

During apoptosis, ICE family members such as caspase-3 and -7 cleave PARP to yield an 85 kDa and a 25 kDa fragment. PARP cleavage is considered to be one of the classical characteristics of apoptosis. Antibodies to the 85 kDa fragment of cleaved PARP or to caspase-3 both serve as markers for apoptotic cells that can be monitored using immunofluorescence.

Multiparameter apoptosis kits are formulated to allow the determination of live and dead cells in addition to the caspase status.

  Detecting apoptosis with CellEvent™ Caspase-3/7 Green detection reagent
Multiplex imaging of U2OS cells using CellEvent™ Caspase-3/7 Green detection reagent to detect apoptosis.

Caspase Assays in Microplates

  • Rhodamine 110-derived caspase substrates are available with a variety of enzyme specificities.
  • Real-time live-cell assays
  • Optimized kits for cell lysates
  • Individual substrates with choice of specificity

Life Technologies offers microplate assays for caspase activity that can be performed on whole cells or on cell lysates. 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.

Microplate kits for caspase-3 measurement provide a choice of substrates with different wavelength emissions: Rhodamine 110 or AMCA can be used to report activity.

  Dose response curve with CellEvent™ Caspase-3/7 Green Detection Reagent.
Dose response curve with CellEvent™ Caspase-3/7 Green Detection Reagent.


Selection Guides for Caspase Assays

Cysteine-dependent aspartate-directed proteases (caspases) play an essential role in different stages of apoptosis. Caspase assays are available for live or formalin-fixed cells. A real-time assay and no-wash protocol is particularly useful in allowing assay multiplexing with delicate apoptotic cells.

 
Readout No-wash assay to monitor ongoing caspase activity. Fluorescence intensity increases with caspase activity.
Caspase 3, 7
Standard filter set FITC
Ex/Em (nm) 502/530
Bibliography Citations
Fluorescence microscope Yes
HCS Yes
HTS Yes
Live cells Yes
Fixed cells No
Fixable Yes
Format 100 µL
Cat. No. C10423

Assay kits based on signal retention require washing and provide a snapshot of caspase activity. Each kit format provides simultaneous detection of apoptotic cells (based on caspase activity), live cells, and dead cells. Immunofluorescence protocols permit staining in fixed cells and tissue sections.

  PARP FITC Apoptosis Kit
Readout Assay kits based on signal retention require washing and provide a snapshot of caspase activity. Each kit format provides simultaneous detection of apoptotic cells (based on caspase activity), live cells, and dead cells. Marker for detecting apoptotic cells.
Standard filter sets DAPI FITC TRITC DAPI FITC Texas Red® FITC
Fluorophore Hoechst 33342 SYTOX® Green FMK Hoechst 33342 FAM Propidium iodide FITC
Ex/Em (nm) 350/461 504/523 550/595 350/461 488/530 535/617 490/525
Bibliography Citations Citations
Fluorescence microscope Yes Yes Yes
HCS Yes Yes Yes
HTS      
Live cells Yes Yes No
Fixed cells No No Yes
Fixable Yes Yes Yes
Caspase Poly-caspase 3, 7 Poly-caspase 8 3, 7 3, 7
Format 25-assay kit 25-assay kit 25-assay kit 25-assay kit 25-assay kit 1 kit, 100 tests
Cat. No. I35101 I35102 I35104 I35105 I35106 AHM2011

Cysteine-dependent aspartate-directed proteases (caspases) play an essential role in different stages of apoptosis. Assays of cell extracts can provide convenient measures of caspase activity for large numbers of samples.

 
ReadoutThese kits can be used to continuously measure the activity of caspase-3 in cell extracts and purified enzyme preparations, using a fluorescence microplate reader.
Conjugated dyeRhodamine 110AMCRhodamine 110
Ex/Em (nm)496/520342/441496/520
BibliographyCitations
Fluorescence microscope   
HCS   
HTSYesYesYes
Selective caspase detectionCaspase-3Caspase-3Caspase-3
Format500-assay kit500-assay kit96-assay kit in preformatted plate
Cat. No.E13184E13183R35100

Caspase substrates and antibody reagents can be incorporated into assays for imaging or screening protocols.

  CASP3 [D175] Monoclonal Antibody (clone 9H19L2), ABfinity™ Recombinant
ReadoutThe rhodamine 110 (R110)-derived caspase substrates are cleaved by the protease, and converted first to the fluorescent monoamide and then to R110, with a further increase in fluorescence.The substrates can be used to continuously measure enzyme activity in cell extracts and purified enzyme preparations using a fluorescence microplate reader.Anti-CASP3 enables caspase detection in fixed and permeabilized cells.
Conjugated dyeRhodamine 110NA (unconjugated rabbit IgG)
Ex/Em (nm)498/521
BibliographyCitationsCitations
Fluorescence microscope   Yes
HCS   Yes
HTSYesYesYes 
Live cells   No
Fixed cells   Yes
Fixable   Yes
Selective caspase detectionGeneral caspase reagentCaspase-1, caspase-4Caspase-6Caspase-3
Format1 mg2 mg2 mg100 µg
Cat. No.R22122R33750R33754700182