Fluorescence-based cell viability screening assays using water-soluble oxygen probes

Area of Science:

• Biotechnology
• Cell Biology
• Assay Development

Background:

• Cell viability assays are crucial for drug discovery and toxicology.
• Existing microplate-based assays can be complex, costly, or lack sensitivity.
• Monitoring cellular respiration offers a direct measure of metabolic activity and cell health.

Purpose of the Study:

• To develop a simple, cost-effective, and robust luminescence-based assay for screening mammalian cell viability.
• To utilize a phosphorescent oxygen probe to monitor cell respiration in a high-throughput microplate format.
• To correlate oxygen consumption rates with cell number, viability, and drug responses.

Main Methods:

• A water-soluble phosphorescent oxygen probe was added to mammalian cell cultures in 96-well plates.
• Cellular oxygen consumption was measured by monitoring changes in phosphorescence intensity and lifetime.
• A mineral oil layer was applied to samples to facilitate oxygen consumption analysis.
• Kinetic changes in phosphorescence were analyzed using prompt or time-resolved fluorescence plate readers.

Main Results:

• The assay successfully monitored oxygen consumption rates in mammalian cells.
• Oxygen uptake rates were correlated with cell numbers (10^5 to 10^7 cells/mL).
• The assay demonstrated sensitivity to cell viability and drug/effector actions.

Conclusions:

• A simple, noninvasive, and cost-effective luminescence assay for cell viability screening was established.
• The assay is compatible with existing instrumentation and suitable for high-throughput analysis.
• This method provides a robust alternative to existing microplate-based cell viability assays.