The fluorescence decay in fluorescence lifetime imaging (FLIM) is typically fitted to a multi-exponential model with discrete lifetimes. The interaction between fluorophores in heterogeneous samples (e.g. biological tissue) can, however, produce complex decay characteristics that do not correspond to such models. Although they appear to provide a better fit to fluorescence decay data than the assumption of a mono-exponential decay, the assumption of multiple discrete components is essentially arbitrary and often erroneous. The stretched exponential function (StrEF) describes fluorescence decay profiles using a continuous lifetime distribution as has been reported for tryptophan, being one of the main fluorophores in tissue. We have demonstrated that this model represents our time-domain FLIM data better than multi-exponential discrete decay components, yielding excellent contrast in tissue discrimination without compromising the goodness of fit, and it significantly decreases the required processing time. In addition, the stretched exponential decay model can provide a direct measure of the sample heterogeneity and the resulting heterogeneity map can reveal subtle tissue differences that other models fail to show.
We report a whole-field fluorescence imaging microscope that combines 3-D spatial resolution by optical sectioning, using structured illumination, with fluorescence lifetime imaging and spectrally-resolved imaging. We show the potential of this technique in the elimination of common artefacts in fluorescence lifetime imaging and apply it to study the dependence of the lifetime on the emission wavelength in biological tissue.
We present potential biomedical applications for a diode- pumped ultrafast Cr:LiSGAF oscillator-amplifier system. A whole-field fluorescence lifetime imaging system has been demonstrated for the first time using such a laser system. Fluorescence lifetime imaging of unstained biological tissue in vitro using this instrument has shown contrast between different tissue constituents. Initial results of applying this laser system to the ablation of glass are also presented.