Summary
The initial steps toward optical detection and spectroscopy of single molecules arose out of the study of spectral hole-burning in inhomogeneously broadened optical absorption profiles of molecular impurities in solids at low temperatures. Spectral signatures relating to the fluctuations of the number of molecules in resonance led to the attainment of the single-molecule limit in 1989. In the early 1990s, many fascinating physical effects were observed for individual molecules such as spectral diffusion, optical switching, vibrational spectra, and magnetic resonance of a single molecular spin. Since the mid-1990s when experiments moved to room temperature, a wide variety of biophysical effects may be explored, and a number of physical phenomena from the low temperature studies have analogs at high temperature. Recent advances worldwide cover a huge range, from in vitro studies of enzymes, proteins, and oligonucleotides, to observations in real time of a single protein performing a specific function inside a living cell. Because each single fluorophore acts a light source roughly 1 nm in size, microscopic observation of individual fluorophores leads naturally to localization beyond the optical diffraction limit. Combining this with active optical control of the number of emitting molecules leads to superresolution imaging, a new frontier for optical microscopy beyond the optical diffraction limit and for chemical design of photoswitchable fluorescent labels. Finally, to study one molecule in aqueous solution without surface perturbations, a new electrokinetic trap is described (the ABEL trap) which can trap single small biomolecules without the need for large dielectric beads.
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Acknowledgements
The author warmly thanks many members of the Moerner single-molecule spectroscopy group for their crucial contributions to the work reported here, in particular: W. P. Ambrose, Th. Basché, T. P. Carter, L. Kador, J. Köhler, D. J. Norris, and T. Plakhotnik (low-temperature studies); J. Biteen, S. Brasselet, A. E. Cohen, N. R. Conley, J. Deich, R. M. Dickson, S.-Y. Kim, B. Lounis, S. Y. Nishimura, E. J. G. Peterman, M. Vrljic, and K. Willets. (room-temperature studies) Stimulating collaborations with the groups of H. M. McConnell, M. Orrit, J. Schmidt, L. Shapiro, R. J. Twieg, and U. P. Wild are gratefully acknowledged. This work has been supported in part by prior grants from the U. S. Office of Naval Research and the National Science Foundation, and by current grants from the National Institutes of Health Nos. 1P20-HG003638, 1R21-RR023149, and 1R01-GM085437, from the Department of Energy Grant No. DE-FG02–07ER15892, and from the National Science Foundation Grant No. DMR-0507296.
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Moerner, W.E. (2010). Single-Molecule Optical Spectroscopy and Imaging: From Early Steps to Recent Advances. In: Gräslund, A., Rigler, R., Widengren, J. (eds) Single Molecule Spectroscopy in Chemistry, Physics and Biology. Springer Series in Chemical Physics, vol 96. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-02597-6_2
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