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Visualization of the Microtubules of Glutaraldehyde-Fixed Cells by Reflection-Enhanced Backscatter Confocal Microscopy

Published online by Cambridge University Press:  09 December 2005

Charles H. Keith  and
Mark A. Farmer
Charles H. Keith
Affiliation:
Department of Cellular Biology, The University of Georgia, 724 Biological Sciences Building, Athens, GA 30602
Mark A. Farmer
Affiliation:
Department of Cellular Biology, The University of Georgia, 724 Biological Sciences Building, Athens, GA 30602 Center for Advanced Ultrastructural Research, The University of Georgia, 151 Barrow Hall, Athens, GA 30602
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Abstract

Performing reflection-mode (backscatter-mode) confocal microscopy on cells growing on reflective substrates gives images that have improved contrast and are more easily interpreted than standard reflection-mode confocal micrographs (Keith et al., 1998). However, a number of factors degrade the quality of images taken with the highest-resolution microscope objectives in this technique. We here describe modifications to reflection-enhanced backscatter confocal microscopy that (partially) overcome these factors. With these modifications of the technique, it is possible to visualize structures the size—and refractility—of individual microtubules in intact cells. Additionally, we demonstrate that this technique, in common with fluorescence techniques such as standing wave widefield fluorescence microscopy and 4-Pi confocal microscopy, offers improved resolution in the Z-direction.

Keywords

imaginghyperresolutionopaque substrates
Type
BIOLOGICAL APPLICATIONS
Information
Microscopy and Microanalysis , Volume 12 , Issue 2 , April 2006 , pp. 113 - 123
Copyright
© 2006 Microscopy Society of America

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