Nature Methods 2, 920 - 931 (2005)
Published online: 18 November 2005; | doi:10.1038/nmeth815
Optical sectioning microscopyJosé-Angel Conchello1, 2
& Jeff W Lichtman31
Molecular, Cell, and Developmental Biology, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma 73104, USA. 2
Program in Biomedical Engineering, University of Oklahoma, Norman, Oklahoma 73019, USA. 3
Molecular and Cell Biology Department, Harvard University, Cambridge, Massachusetts 02138, USA.
Correspondence should be addressed to José-Angel Conchello jose-conchello@omrf.ouhsc.edu Confocal scanning microscopy, a form of optical sectioning microscopy, has radically transformed optical imaging in biology. These devices provide a powerful means to eliminate from images the background caused by out-of-focus light and scatter. Confocal techniques can also improve the resolution of a light microscope image beyond what is achievable with widefield fluorescence microscopy. The quality of the images obtained, however, depends on the user's familiarity with the optical and fluorescence concepts that underlie this approach. We describe the core concepts of confocal microscopes and important variables that adversely affect confocal images. We also discuss data-processing methods for confocal microscopy and computational optical sectioning techniques that can perform optical sectioning without a confocal microscope.
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