Using Becker & Hickl TCSPC FLIM systems with fast TCSPC modules and fast detectors, we found ultra-fast fluorescence decay components in biological material, such as mushroom spores, pollen grains, plant tissue, and malignant melanoma. The component decay times are on the order of 10 to 50 ps, the amplitudes range from about 60% to more than 99%. In general, the shortest lifetimes and the largest amplitudes of the fast component were found in strongly coloured material, such as black mushroom spores and melanoma tissue. However, the lifetime not always correlates with the colour. In particular, a fast decay component was not found in tissue from basal cell papilloma, although it is dark brown. This may open a way to identify melanoma tissue and melanoma cells, and thus provide a new tool to investigate melanoma progression.
Ophthalmic imaging by fluorescence techniques is a tool which gets more and more established in eye disease diagnosis and research. All type of clinical imaging is usually restricted to the use of endogenous fluorophores present in the tissue. The excitation and emission spectra of these fluorophores are overlapping and poorly defined. Moreover, the apparent spectra are changed by variation in the relative concentration of fluorophores and by absorbers present in the tissue. Intensity images, even those with spectral resolution, therefore deliver very limited information on the state of the tissue.
A considerable improvement in the field of retinal imaging is obtained by using fluorescence lifetime imaging ophthalmoscopy (FLIO). The fluorescence lifetime measured by TCSPC is independent of the concentration, and enables the possibility to measure even the weak retinal autofluorescence. Moreover, it delivers direct information on the configuration of endogenous fluorophores, on binding to proteins or lipids, on the redox state, and on other metabolic parameters.
We will describe the technical problems of FLIO data and their solutions, demonstrate the performance of existing systems in ophthalmology and present some results.
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