Chapter title |
Does Super-Resolution Fluorescence Microscopy Obsolete Previous Microscopic Approaches to Protein Co-localization?
|
---|---|
Chapter number | 19 |
Book title |
Membrane Trafficking
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Published in |
Methods in molecular biology, January 2015
|
DOI | 10.1007/978-1-4939-2309-0_19 |
Pubmed ID | |
Book ISBNs |
978-1-4939-2308-3, 978-1-4939-2309-0
|
Authors |
Laura MacDonald, Giulia Baldini, Brian Storrie, MacDonald, Laura, Baldini, Giulia, Storrie, Brian |
Abstract |
Conventional microscopy techniques, namely, the confocal microscope or deconvolution processes, are resolution limited to approximately 200-250 nm by the diffraction properties of light as developed by Ernst Abbe in 1873. This diffraction limit is appreciably above the size of most multi-protein complexes, which are typically 20-50 nm in diameter. In the mid-2000s, biophysicists moved beyond the diffraction barrier by structuring the illumination pattern and then applying mathematical principles and algorithms to allow a resolution of approximately 100 nm, sufficient to address protein subcellular co-localization questions. This "breaking" of the diffraction barrier, affording resolution beyond 200 nm, is termed super-resolution microscopy. More recent approaches include single-molecule localization (such as photoactivated localization microscopy (PALM)/stochastic optical reconstruction microscopy (STORM)) and point spread function engineering (such as stimulated emission depletion (STED) microscopy). In this review, we explain basic principles behind currently commercialized super-resolution setups and address advantages and considerations in applying these techniques to protein co-localization in biological systems. |
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