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

by Winter, Peter W , Waterman, Clare M , Senseney, Justin , York, Andrew G , Wawrzusin, Peter , Wu, Yicong , Fischer, Robert S , Santella, Anthony , McAuliffe, Matthew , Colón-Ramos, Daniel A , Bao, Zhirong , Christensen, Ryan , Shroff, Hari

in 631/136/334/1582/712 / 631/1647/328/2237 / 631/80/128/1653 / Agriculture / Animals / Bioinformatics / Biomedical Engineering/Biotechnology / Biomedicine / Biotechnology / Brain - anatomy & histology / Brain - growth & development / Brain - ultrastructure / Caenorhabditis elegans / Caenorhabditis elegans - embryology / Cell culture / Cell research / Cell Tracking - methods / Embryonic growth stage / Equipment and supplies / Fourth dimension / Human Umbilical Vein Endothelial Cells / Humans / Imaging systems / Imaging, Three-Dimensional - methods / Innovations / Life Sciences / Lighting / Microscopes / Microscopy / Microscopy - instrumentation / Microscopy - methods / Nematoda / Observations / Optical microscopes / Optical properties / Photobleaching / Spatial light modulators / Volumetric analysis

2013

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

2013

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

2013

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Journal Article

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

Winter, Peter W,

Waterman, Clare M,

Senseney, Justin,

York, Andrew G,

Wawrzusin, Peter,

Wu, Yicong,

Fischer, Robert S,

Santella, Anthony,

McAuliffe, Matthew,

Colón-Ramos, Daniel A,

Bao, Zhirong,

Christensen, Ryan,

Shroff, Hari

2013

Overview

A dual-view light sheet microscope allows high-speed imaging with an isotropic spatial resolution 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.

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Publisher

Nature Publishing Group UK,Nature Publishing Group

Subject

/ Animals

/ Biomedical Engineering/Biotechnology