Fast objective coupled planar illumination microscopy

Nat Commun. 2019 Oct 2;10(1):4483. doi: 10.1038/s41467-019-12340-0.

Abstract

Among optical imaging techniques light sheet fluorescence microscopy is one of the most attractive for capturing high-speed biological dynamics unfolding in three dimensions. The technique is potentially millions of times faster than point-scanning techniques such as two-photon microscopy. However light sheet microscopes are limited by volume scanning rate and/or camera speed. We present speed-optimized Objective Coupled Planar Illumination (OCPI) microscopy, a fast light sheet technique that avoids compromising image quality or photon efficiency. Our fast scan system supports 40 Hz imaging of 700 μm-thick volumes if camera speed is sufficient. We also address the camera speed limitation by introducing Distributed Planar Imaging (DPI), a scaleable technique that parallelizes image acquisition across cameras. Finally, we demonstrate fast calcium imaging of the larval zebrafish brain and find a heartbeat-induced artifact, removable when the imaging rate exceeds 15 Hz. These advances extend the reach of fluorescence microscopy for monitoring fast processes in large volumes.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Brain / diagnostic imaging*
  • Diagnostic Imaging / instrumentation*
  • Diagnostic Imaging / methods
  • Image Processing, Computer-Assisted / instrumentation*
  • Image Processing, Computer-Assisted / methods
  • Larva
  • Luminescent Measurements / instrumentation*
  • Luminescent Measurements / methods
  • Microscopy / instrumentation*
  • Microscopy / methods
  • Microscopy, Confocal / instrumentation
  • Microscopy, Confocal / methods
  • Microscopy, Fluorescence / instrumentation
  • Microscopy, Fluorescence / methods
  • Microscopy, Fluorescence, Multiphoton / instrumentation
  • Microscopy, Fluorescence, Multiphoton / methods
  • Reproducibility of Results
  • Zebrafish