Single-molecule fluorescence methods, particularly single-molecule FRET (fluorescence resonance energy transfer), have provided novel insights into the structure, interactions and dynamics of biological systems. ALEX (alternating-laser excitation) spectroscopy is a new method that extends single-molecule FRET by providing simultaneous information about structure and stoichiometry; this new information allows the detection of interactions in the absence of FRET and extends the dynamic range of distance measurements that are accessible through FRET. In the present article, we discuss combinations of ALEX with confocal microscopy for studying in-solution and in-gel molecules; we also discuss combining ALEX with TIRF (total internal reflection fluorescence) for studying surface-immobilized molecules. We also highlight applications of ALEX to the study of protein–nucleic acid interactions.
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August 2008
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Conference Article|
July 22 2008
Red light, green light: probing single molecules using alternating-laser excitation
Yusdi Santoso;
Yusdi Santoso
1Department of Physics and Interdisciplinary Research Collaboration in Bionanotechnology, Clarendon Laboratory, University of Oxford, Parks Road, Oxford OX1 3PU, U.K.
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Ling Chin Hwang;
Ling Chin Hwang
1Department of Physics and Interdisciplinary Research Collaboration in Bionanotechnology, Clarendon Laboratory, University of Oxford, Parks Road, Oxford OX1 3PU, U.K.
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Ludovic Le Reste;
Ludovic Le Reste
1Department of Physics and Interdisciplinary Research Collaboration in Bionanotechnology, Clarendon Laboratory, University of Oxford, Parks Road, Oxford OX1 3PU, U.K.
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Achillefs N. Kapanidis
Achillefs N. Kapanidis
1
1Department of Physics and Interdisciplinary Research Collaboration in Bionanotechnology, Clarendon Laboratory, University of Oxford, Parks Road, Oxford OX1 3PU, U.K.
1To whom correspondence should be addressed (email [email protected]).
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Publisher: Portland Press Ltd
Received:
April 07 2008
Online ISSN: 1470-8752
Print ISSN: 0300-5127
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem Soc Trans (2008) 36 (4): 738–744.
Article history
Received:
April 07 2008
Citation
Yusdi Santoso, Ling Chin Hwang, Ludovic Le Reste, Achillefs N. Kapanidis; Red light, green light: probing single molecules using alternating-laser excitation. Biochem Soc Trans 1 August 2008; 36 (4): 738–744. doi: https://doi.org/10.1042/BST0360738
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