Please use this identifier to cite or link to this item:
Title: Programmable Illumination and High-Speed, Multi- Wavelength, Confocal Microscopy Using a Digital Micromirror
Authors: Martial, Franck P.
Hartell, Nicholas A.
First Published: 24-Aug-2012
Publisher: Public Library of Science
Citation: PLoS One, 2012, 7 (8), e43942
Abstract: Confocal microscopy is routinely used for high-resolution fluorescence imaging of biological specimens. Most standard confocal systems scan a laser across a specimen and collect emitted light passing through a single pinhole to produce an optical section of the sample. Sequential scanning on a point-by-point basis limits the speed of image acquisition and even the fastest commercial instruments struggle to resolve the temporal dynamics of rapid cellular events such as calcium signals. Various approaches have been introduced that increase the speed of confocal imaging. Nipkov disk microscopes, for example, use arrays of pinholes or slits on a spinning disk to achieve parallel scanning which significantly increases the speed of acquisition. Here we report the development of a microscope module that utilises a digital micromirror device as a spatial light modulator to provide programmable confocal optical sectioning with a single camera, at high spatial and axial resolution at speeds limited by the frame rate of the camera. The digital micromirror acts as a solid state Nipkov disk but with the added ability to change the pinholes size and separation and to control the light intensity on a mirror-by-mirror basis. The use of an arrangement of concave and convex mirrors in the emission pathway instead of lenses overcomes the astigmatism inherent with DMD devices, increases light collection efficiency and ensures image collection is achromatic so that images are perfectly aligned at different wavelengths. Combined with non-laser light sources, this allows low cost, high-speed, multi-wavelength image acquisition without the need for complex wavelength-dependent image alignment. The micromirror can also be used for programmable illumination allowing spatially defined photoactivation of fluorescent proteins. We demonstrate the use of this system for high-speed calcium imaging using both a single wavelength calcium indicator and a genetically encoded, ratiometric, calcium sensor.
DOI Link: 10.1371/journal.pone.0043942
ISSN: 1932-6203
eISSN: 1932-6203
Version: Publisher Version
Status: Peer-reviewed
Type: Journal Article
Rights: Copyright © 2012 Martial, Hartell. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Appears in Collections:Published Articles, Dept. of Cell Physiology and Pharmacology

Files in This Item:
File Description SizeFormat 
journal pone 00439421.pdfPublished (publisher PDF)742 kBAdobe PDFView/Open

Items in LRA are protected by copyright, with all rights reserved, unless otherwise indicated.