Line excitation array detection fluorescence microscopy at 0.8 million frames per second

Nat Commun. 2018 Oct 29;9(1):4499. doi: 10.1038/s41467-018-06775-0.

Abstract

Three-dimensional, fluorescence imaging methods with ~1 MHz frame rates are needed for high-speed, blur-free flow cytometry and capturing volumetric neuronal activity. The frame rates of current imaging methods are limited to kHz by the photon budget, slow camera readout, and/or slow laser beam scanners. Here, we present line excitation array detection (LEAD) fluorescence microscopy, a high-speed imaging method capable of providing 0.8 million frames per second. The method performs 0.8 MHz line-scanning of an excitation laser beam using a chirped signal-driven longitudinal acousto-optic deflector to create a virtual light-sheet, and images the field-of-view with a linear photomultiplier tube array to generate a 66 × 14 pixel frame each scan cycle. We implement LEAD microscopy as a blur-free flow cytometer for Caenorhabditis elegans moving at 1 m s-1 with 3.5-µm resolution and signal-to-background ratios >200. Signal-to-noise measurements indicate future LEAD fluorescence microscopes can reach higher resolutions and pixels per frame without compromising frame rates.

Publication types

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

MeSH terms

  • Animals
  • Caenorhabditis elegans / drug effects
  • Dimethyl Sulfoxide / pharmacology
  • Flow Cytometry / instrumentation
  • Flow Cytometry / methods
  • Imaging, Three-Dimensional / instrumentation*
  • Imaging, Three-Dimensional / methods*
  • Microscopy, Fluorescence / instrumentation*
  • Microscopy, Fluorescence / methods*
  • Models, Biological
  • Neurons
  • Optics and Photonics / instrumentation
  • Optics and Photonics / methods
  • Peptides
  • Photons
  • Protein Aggregation, Pathological / diagnostic imaging
  • Sensitivity and Specificity

Substances

  • Peptides
  • polyglutamine
  • Dimethyl Sulfoxide