Live-cell Imaging of Single-Cell Arrays (LISCA) - a Versatile Technique to Quantify Cellular Kinetics

Area of Science:

• Cellular and Molecular Biology
• Biophysics
• Microscopy and Imaging

Background:

• Single-cell fluorescence time-course acquisition is challenging due to cell motility and shape variability.
• Adhesive micro-arrays offer a solution by standardizing cellular conditions and simplifying image analysis.

Purpose of the Study:

• To detail the experimental workflow for Live-cell Imaging of Single-Cell Arrays (LISCA).
• To establish a method for high-throughput, parallel monitoring of single-cell fluorescence.
• To analyze mRNA expression kinetics and degradation rates.

Main Methods:

• Utilizing adhesive micro-arrays with micropatterned surfaces for cell culture.
• Employing scanning time-lapse microscopy to capture fluorescence signals from individual cells.
• Developing automated image processing software to generate single-cell fluorescence time courses from eGFP expression.

Main Results:

• LISCA successfully captures time courses of enhanced green fluorescent protein (eGFP) expression in hundreds of cells simultaneously.
• A simple kinetic translation model accurately describes eGFP expression kinetics, enabling determination of mRNA expression and degradation rates.
• The method proves effective for analyzing single-cell fluorescence dynamics.

Conclusions:

• LISCA is a robust and high-throughput method for live-cell imaging of single-cell fluorescence dynamics.
• The technique allows for quantitative analysis of mRNA kinetics at the single-cell level.
• LISCA has potential applications in studying signaling pathways and apoptosis.