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We report the cell biological applications of a recently developedmultiphoton fluorescence lifetime imaging microscopy system using a streakcamera (StreakFLIM). The system was calibrated with standard fluorophorespecimens and was shown to have high accuracy and reproducibility. Wedemonstrate the applicability of this instrument in living cells for measuringthe effects of protein targeting and point mutations in the protein sequencewhich are not obtainable in conventional intensity based fluorescencemicroscopy methods. We discuss the relevance of such time resolved informationin quantitative energy transfer microscopy and in measurement of the parameterscharacterizing intracellular physiology

Probing subtle fluorescence dynamics in cellular proteins by streak camera based fluorescence lifetime imaging microscopy