Affiliation:
1. Department of Mathematics and Center for Cellular Nanoanalytics , Osnabrück University , Barbarastrasse 113, D-49076 Osnabruck , Germany
Abstract
Abstract
Classical fluorescence microscopy is a powerful technique to image biological specimen under close-to-native conditions, but light diffraction limits its optical resolution to 200–300 nm-two orders of magnitude worse than the size of biomolecules. Assuming single fluorescent emitters, the final image of the optical system can be described by a convolution with the point spread function (PSF) smearing out details below the size of the PSF. In mathematical terms, fluorescence microscopy produces bandlimited space-continuous images that can be recovered from their spatial samples under the conditions of the classical Shannon-Nyquist theorem. During the past two decades, several single molecule localization techniques have been established and these allow for the determination of molecular positions with sub-pixel accuracy. Without noise, single emitter positions can be recovered precisely – no matter how close they are. We review recent work on the computational resolution limit with a sharp phase transition between two scenarios: 1) where emitters are well-separated with respect to the bandlimit and can be recovered up to the noise level and 2) closely distributed emitters which results in a strong noise amplification in the worst case. We close by discussing additional pitfalls using single molecule localization techniques based on structured illumination.
Funder
Volkswagen Foundation
Deutsche Forschungsgemeinschaft
Subject
Clinical Biochemistry,Molecular Biology,Biochemistry
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