title: Digitally synthesized beat frequency-multiplexed fluorescence lifetime spectroscopy.
abstract: Frequency domain fluorescence lifetime imaging is a powerful technique that enables the observation of subtle changes in the molecular environment of a fluorescent probe. This technique works by measuring the phase delay between the optical emission and excitation of fluorophores as a function of modulation frequency. However, high-resolution measurements are time consuming, as the excitation modulation frequency must be swept, and faster low-resolution measurements at a single frequency are prone to large errors. Here, we present a low cost optical system for applications in real-time confocal lifetime imaging, which measures the phase vs. frequency spectrum without sweeping. Deemed Lifetime Imaging using Frequency-multiplexed Excitation (LIFE), this technique uses a digitally-synthesized radio frequency comb to drive an acousto-optic deflector, operated in a cat's-eye configuration, to produce a single laser excitation beam modulated at multiple beat frequencies. We demonstrate simultaneous fluorescence lifetime measurements at 10 frequencies over a bandwidth of 48 MHz, enabling high speed frequency domain lifetime analysis of single- and multi-component sample mixtures.
authors.affiliation: Departments of Electrical Engineering, University of California, Los Angeles, CA 90095, USA.
authors.forename: Jacky C K
authors.initials: JC
authors.lastname: Chan
authors.affiliation: Departments of Electrical Engineering, University of California, Los Angeles, CA 90095, USA ; Department of Bioengineering, University of California, Los Angeles, CA 90095, USA.
authors.forename: Eric D
authors.initials: ED
authors.lastname: Diebold
authors.affiliation: Departments of Electrical Engineering, University of California, Los Angeles, CA 90095, USA.
authors.forename: Brandon W
authors.initials: BW
authors.lastname: Buckley
authors.affiliation: Departments of Electrical Engineering, University of California, Los Angeles, CA 90095, USA.
authors.forename: Sien
authors.initials: S
authors.lastname: Mao
authors.affiliation: Departments of Electrical Engineering, University of California, Los Angeles, CA 90095, USA.
authors.forename: Najva
authors.initials: N
authors.lastname: Akbari
authors.affiliation: Departments of Electrical Engineering, University of California, Los Angeles, CA 90095, USA ; Department of Bioengineering, University of California, Los Angeles, CA 90095, USA ; Department of Surgery, David Geffen School of Medicine, University of California, Los Angeles, CA 90095, USA.
authors.forename: Bahram
authors.initials: B
authors.lastname: Jalali
keywords: (170.1065) Acousto-optics; (170.1790) Confocal microscopy; (170.2520) Fluorescence microscopy; (170.3650) Lifetime-based sensing