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Updated: Nov 4, 2025
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Autonomously Bioluminescent Mammalian Cells for Continuous and Real-time Monitoring of Cytotoxicity
Published on: October 28, 2013
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A Real-Time, Bioluminescent Apoptosis Assay.
Kevin R Kupcho1, Andrew L Niles2
1Promega, Madison, WI, USA.
Methods in Molecular Biology (Clifton, N.J.)
|May 25, 2021
Summary
This novel bioluminescent assay tracks phosphatidylserine (PS) exposure in real-time, overcoming timing issues in apoptosis studies. It offers kinetic resolution of cell death, from early to late phases, for various inducers.
Area of Science:
- Cell Biology
- Biochemistry
- Assay Development
Background:
- Traditional apoptosis assays face timing challenges with unknown inducer potentials.
- Monitoring phosphatidylserine (PS) translocation is crucial for detecting early apoptosis.
- Existing methods often require multiple steps and lack real-time kinetic data.
Purpose of the Study:
- To present a real-time, bioluminescent assay for continuous apoptosis monitoring.
- To overcome the 'timing conundrum' associated with traditional apoptosis detection.
- To provide kinetic resolution of dose- and agent-dependent apoptotic responses.
Main Methods:
- Utilized a homogenous, no-wash, plate-based assay.
- Employed annexin V fusion proteins with complementing NanoBiT™ luciferase subunits.
- Incorporated a time-released luciferase substrate and a fluorescent membrane integrity reagent.
Main Results:
- The assay continuously reports phosphatidylserine (PS) translocation during apoptosis.
- Luminescence signal is proportional to PS exposure, indicating early apoptotic events.
- Fluorescence intensity correlates with secondary necrosis, allowing late-phase detection.
Conclusions:
- This bioluminescent technique offers exquisite kinetic resolution of apoptosis.
- The assay circumvents timing issues, enabling precise analysis of cell death kinetics.
- Compatible with additional reagents for orthogonal cell health measurements.
