We present a technique that records transient changes in the fluorescence lifetime of a sample with spatial resolution along a one-dimensional scan. The technique is based on scanning the sample with a high-frequency pulsed laser beam, detecting single photons of the fluorescence light, and building up a photon distribution over the distance along the scan, the arrival times of the photons after the excitation pulses and the time after a stimulation of the sample. The maximum resolution at which lifetime changes can be recorded is given by the line scan period. Transient lifetime effects can thus be resolved at a resolution of about one millisecond. We demonstrate the technique for recording photochemical and nonphotochemical chlorophyll transients in plants and transient changes in free Ca(2+) in cultured neurons.
Keywords: Ca2+ imaging; FLIM; FLITS; TCSPC; chlorophyll transients; fluorescence lifetime.
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