Spatially isotropic four-dimensional imaging with dual-view plane illumination microscopy.

Imagen de Daniel Alfonso Colón-Ramos
PDF versionPDF version
TítuloSpatially isotropic four-dimensional imaging with dual-view plane illumination microscopy.
Publication TypeJournal Article
Year of Publication2013
AutoresWu, Y, Wawrzusin, P, Senseney, J, Fischer, RS, Christensen, R, Santella, A, York, AG, Winter, PW, Waterman, CM, Bao, Z, Colón-Ramos, DA, McAuliffe, M, Shroff, H
JournalNat Biotechnol
Volume31
Issue11
Pagination1032-8
Date Published2013 Nov
ISSN1546-1696
Palabras claveAnimals, Brain, Caenorhabditis elegans, Cell Tracking, Human Umbilical Vein Endothelial Cells, Humans, Imaging, Three-Dimensional, Lighting, Microscopy, Photobleaching
Abstract

Optimal four-dimensional imaging requires high spatial resolution in all dimensions, high speed and minimal photobleaching and damage. We developed a dual-view, plane illumination microscope with improved spatiotemporal resolution by switching illumination and detection between two perpendicular objectives in an alternating duty cycle. Computationally fusing the resulting volumetric views provides an isotropic resolution of 330 nm. As the sample is stationary and only two views are required, we achieve an imaging speed of 200 images/s (i.e., 0.5 s for a 50-plane volume). Unlike spinning-disk confocal or Bessel beam methods, which illuminate the sample outside the focal plane, we maintain high spatiotemporal resolution over hundreds of volumes with negligible photobleaching. To illustrate the ability of our method to study biological systems that require high-speed volumetric visualization and/or low photobleaching, we describe microtubule tracking in live cells, nuclear imaging over 14 h during nematode embryogenesis and imaging of neural wiring during Caenorhabditis elegans brain development over 5 h.

DOI10.1038/nbt.2713
Alternate JournalNat. Biotechnol.
PubMed ID24108093
PubMed Central IDPMC4105320
Grant ListGM097576 / GM / NIGMS NIH HHS / United States
HD075602 / HD / NICHD NIH HHS / United States
R01 NS076558 / NS / NINDS NIH HHS / United States
R01NS076558 / NS / NINDS NIH HHS / United States
U01 HD075602 / HD / NICHD NIH HHS / United States
U01HD075602 / HD / NICHD NIH HHS / United States