Chapter 6 Flow cytometric fluorescence lifetime measurements

Abstract

Publisher Summary Fluorescence lifetime measurements by flow cytometry (FCM) are important because they yield additional information about fluorophore-cell interactions at the molecular level. An advantage of fluorescence lifetime measurements is that, in some instances, lifetimes can be considered as absolute quantities. There are two general methods for measuring fluorescence lifetimes: time-domain and frequency-domain analysis. In the time-domain method, the sample is excited by a series of short light pulses, and the time evolution of the fluorescence emission is measured directly by time-correlated single photon counting or a high-speed digital storage oscilloscope/sampling. Phase-modulation measurement is a frequency-domain alternative to the timedomain methods. In contrast to pulsed techniques that record the amplitude of the fluorescence decay directly, the phase-modulation method determines the phase and amplitude of the sample fluorescence emission relative to a periodically modulated excitation source.