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Fluorescence Lifetime Imaging of Molecular Rotors in Living Cells
Published on: February 9, 2012
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Digitally synthesized beat frequency-multiplexed fluorescence lifetime spectroscopy.
Jacky C K Chan1, Eric D Diebold2, Brandon W Buckley1
1Departments of Electrical Engineering, University of California, Los Angeles, CA 90095, USA.
Biomedical Optics Express
|January 10, 2015
Summary
This study introduces Lifetime Imaging using Frequency-multiplexed Excitation (LIFE), a novel system for rapid, real-time confocal fluorescence lifetime imaging. LIFE achieves high-speed measurements by analyzing multiple frequencies simultaneously, overcoming limitations of traditional methods.
Area of Science:
- * Optical Imaging
- * Spectroscopy
- * Biophysics
Background:
- * Frequency domain fluorescence lifetime imaging (FD-FLIM) reveals molecular environment changes.
- * Traditional FD-FLIM requires frequency sweeps (time-consuming) or single-frequency measurements (error-prone).
Purpose of the Study:
- * To develop a low-cost optical system for real-time confocal lifetime imaging.
- * To enable high-speed FD-FLIM without sweeping the modulation frequency.
Main Methods:
- * Introduced Lifetime Imaging using Frequency-multiplexed Excitation (LIFE).
- * Utilized a digitally-synthesized radio frequency comb to modulate a laser excitation beam at multiple frequencies.
- * Employed an acousto-optic deflector in a cat's-eye configuration.
Main Results:
- * Demonstrated simultaneous fluorescence lifetime measurements at 10 frequencies across a 48 MHz bandwidth.
- * Achieved high-speed FD-FLIM analysis of complex sample mixtures.
Conclusions:
- * LIFE offers a low-cost, high-speed solution for real-time confocal lifetime imaging.
- * The technique enhances the speed and accuracy of FD-FLIM analysis for various applications.
