Light Microscopy Imaging Facilities

Jeremy Sanderson, University of Sheffield, United Kingdom

Published online: January 2010

DOI: 10.1002/9780470015902.a0022189

Historically core imaging facilities have been associated with electron microscopes. With the very widespread use of fluorescent markers for both fixed and living cells and tissues the light microscope facility has emerged in its own right. Well-run core light microscope imaging facilities are focal points within research institutions. Researchers want to do science, acquire images and manipulate data for publication; for them, the microscope and the computing infrastructure within the imaging facility are merely tools. Staff provide organization, continuity, tuition and knowledge and maintenance of equipment, which encompass the needs of both novice and advanced users. Increasingly, imaging facilities are considered from the outset as a component in the design of a new research building, and must recover the costs of operation. This article discusses the need for optical sectioning, the role of the light microscopy facility and points to consider in its design.

Key Concepts:

  • Fluorescent labelling is widely used in life science to study organelles, structures and proteins of interest within cells and tissues.
  • Fluorescent cells and tissues are self-luminous and light up throughout their thickness. This is often greater than the depth of field (thickness of the in-focus object plane) of the microscope objective, thus most of the image is composed of scattered light and out-of-focus blur which obscures the in-focus detail and drastically reduces the signal-to-noise ratio (SNR).
  • Optical sectioning microscopy ‘cleans up’ the image at each point of focus, so allowing a sharply defined 3D Z-stack to be constructed and analysed.
  • The light microscopes required to acquire multidimensional images over time, and the software packages needed to analyse the data, are both expensive and complex. Core image facilities provide the instrumentation and specialist staff to assist in this process.
  • The design and operation of a core light microscopy imaging facility which is capable of effectively supporting a research institution requires some forethought.

Keywords: imaging facility; optical sectioning; fluorescence; confocal microscopy; deconvolution; facility design

Figure 1. Advantages and disadvantages of fluorescence. PMT, photo-multiplier tube.
Figure 2. Schematic representation for why we need optical sectioning.
Figure 3. Schematic representation of the different types of optical sectioning.
Figure 4. Poster to show facility users which microscope to use.
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Related Sub-Topics:

Techniques in Cell Biology | Optical Microscopy
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Article Title: Light Microscopy Imaging Facilities
 
How to Cite close
Sanderson, Jeremy(Jan 2010) Light Microscopy Imaging Facilities. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0022189]