Metabolic imaging by simultaneous FLIM of NAD(P)H and FAD

Wolfgang Becker; Axel Bergmann; Rodrigo Suarez Ibarrola; Philippe-Fabian Müller; Lukas Braun

doi:10.1117/12.2510132

22 February 2019 Metabolic imaging by simultaneous FLIM of NAD(P)H and FAD

Wolfgang Becker, Axel Bergmann, Rodrigo Suarez Ibarrola, Philippe-Fabian Müller, Lukas Braun

Author Affiliations +

Proceedings Volume 10882, Multiphoton Microscopy in the Biomedical Sciences XIX; 108820B (2019) https://doi.org/10.1117/12.2510132
Event: SPIE BiOS, 2019, San Francisco, California, United States

Abstract

We describe a metabolic-imaging system based on simultaneous recording of lifetime images of NAD(P)H and FAD. The system uses one-photon excitation by ps diode lasers, scanning by galvanometer mirrors, confocal detection, and two parallel TCSPC FLIM recording channels. The two lasers, with wavelengths of 375nm and 405 nm, are multiplexed to alternatingly excite NAD(P)H and FAD. One FLIM channel detects in the emission band of NAD(P)H, the other in the emission band of FAD. The FLIM data are processed by SPCImage data analysis software. For both channels, the data analysis delivers images of the amplitude-weighted lifetime, tm, the component lifetimes, t1 and t2, the amplitudes of the components, a1 and a2, and the amplitude ratio, a1/a2. Moreover, it delivers the fluorescence-lifetime redox ratio (FLIRR), a_2nadh/a_1fad. We demonstrate the performance of the system at the example of human bladder cells. Normal cells and tumor cells were discriminated by the tm images, the a1 images, and the FLIRR images.

Conference Presentation

Citation Download Citation

Wolfgang Becker, Axel Bergmann, Rodrigo Suarez Ibarrola, Philippe-Fabian Müller, and Lukas Braun "Metabolic imaging by simultaneous FLIM of NAD(P)H and FAD", Proc. SPIE 10882, Multiphoton Microscopy in the Biomedical Sciences XIX, 108820B (22 February 2019); https://doi.org/10.1117/12.2510132

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