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Lightsheet fluorescence lifetime imaging microscopy with wide‐field time‐correlated single photon counting
Authors:Liisa M Hirvonen  Jakub Nedbal  Norah Almutairi  Thomas A Phillips  Wolfgang Becker  Thomas Conneely  James Milnes  Susan Cox  Stephen Stürzenbaum  Klaus Suhling
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.image
Keywords:fluorescence lifetime imaging (FLIM)  lightsheet microscopy  microchannel plate (MCP)  SPIM  time‐correlated single photon counting (TCSPC)
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