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High resolution fluorescence microscopy uses the properties of fluorescent markers to overcome the diffraction limit. In contrast to ensemble-based methods (e.g. STED microscopy ), molecules are manipulated individually in SMS (single marker switching) microscopy. In this scheme, single markers are randomly selected from a vast amount of markers being in a dark state and transferred to a bright, detectable state by a stochastic on-switching process. The position of such a single molecule is calculated from its diffraction-limited fluorescence image which is separated spatially and temporally from the spots of other molecules. Here, the localization precision is better than the diffraction limit and scales with √N where N is the number of detected photons.
After the molecule is transferred to a dark state, this process of switching on, reading out and switching off of randomly selected markers is repeated a sufficient number of times; the histogram of all collected marker positions represents the final super resolved SMS image.
Themes within SMS microscopy:
Hell, S. W. (2008):
Egner, A., C. Geisler, C. von Middendorff, H. Bock, D. Wenzel, R. Medda, M. Andresen, A. C. Stiel, S. Jakobs, C. Eggeling, A. Schönle, S. W. Hell (2007):
Geisler, C., A. Schönle, C. von Middendorf, H. Bock, C. Eggeling, A. Egner, S.W. Hell (2007):
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