Fluorescence lifetime imaging (FLIM) of metabolic coenzymes, as NAD(P)H and FAD and oxygen induced phosphorescence lifetime imaging (PLIM) of phosphorescent drugs or delayed FLIM (dFLIM) is now widely accepted to be one of the most important imaging methods for cell metabolism and oxygen. Different algorithms are developed and optimized for simultaneous FLIM/PLIM/dFLIM to get reproducible results. The interaction of light, drug and oxygen during PDT (photodynamic therapy) induces a cellular response, which could be followed by various techniques. The aim of this work was to simultaneously image the cellular response and oxygen consumption in living cells with subcellular resolution.
FLIM of metabolic coenzymes, as NAD(P)H and FAD, is now widely accepted to be one of the most important imaging methods for cell metabolism. However, new algorithms are needed to circumvent various problems and to image cell metabolism and redox state from fluorescence lifetimes. The significance of a metabolic index based on NAD(P)H FLIM will be explained and compared with the fluorescence lifetime induced redox ratio (FLIRR). The importance of FMN will be discussed and the FLIRR approach will be extended. Using a three channel TCSPC system simultaneous metabolic NADH/FAD/FMN FLIM and oxygen PLIM/dFLIM (delayed fluorescence) could be realized.
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