Lightsheet fluorescence lifetime imaging microscopy with wide‐field time‐correlated single photon counting |
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Authors: | Liisa M. Hirvonen,Jakub Nedbal,Norah Almutairi,Thomas A. Phillips,Wolfgang Becker,Thomas Conneely,James Milnes,Susan Cox,Stephen Stü rzenbaum,Klaus Suhling |
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Abstract: | We report on wide‐field time‐correlated single photon counting (TCSPC)‐based fluorescence lifetime imaging microscopy (FLIM) with lightsheet illumination. A pulsed diode laser is used for excitation, and a crossed delay line anode image intensifier, effectively a single‐photon sensitive camera, is used to record the position and arrival time of the photons with picosecond time resolution, combining low illumination intensity of microwatts with wide‐field data collection. We pair this detector with the lightsheet illumination technique, and apply it to 3D FLIM imaging of dye gradients in human cancer cell spheroids, and C. elegans. |
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Keywords: | fluorescence lifetime imaging (FLIM) lightsheet microscopy microchannel plate (MCP) SPIM time‐correlated single photon counting (TCSPC) |
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