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Proceedings Paper

Enhanced simulator software for image validation and interpretation for multimodal localization super-resolution fluorescence microscopy
Author(s): Miklós Erdélyi; József Sinkó; Tamás Gajdos; Tibor Novák
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Paper Abstract

Optical super-resolution techniques such as single molecule localization have become one of the most dynamically developed areas in optical microscopy. These techniques routinely provide images of fixed cells or tissues with sub-diffraction spatial resolution, and can even be applied for live cell imaging under appropriate circumstances. Localization techniques are based on the precise fitting of the point spread functions (PSF) to the measured images of stochastically excited, identical fluorescent molecules. These techniques require controlling the rate between the on, off and the bleached states, keeping the number of active fluorescent molecules at an optimum value, so their diffraction limited images can be detected separately both spatially and temporally. Because of the numerous (and sometimes unknown) parameters, the imaging system can only be handled stochastically. For example, the rotation of the dye molecules obscures the polarization dependent PSF shape, and only an averaged distribution – typically estimated by a Gaussian function – is observed. TestSTORM software was developed to generate image stacks for traditional localization microscopes, where localization meant the precise determination of the spatial position of the molecules. However, additional optical properties (polarization, spectra, etc.) of the emitted photons can be used for further monitoring the chemical and physical properties (viscosity, pH, etc.) of the local environment. The image stack generating program was upgraded by several new features, such as: multicolour, polarization dependent PSF, built-in 3D visualization, structured background. These features make the program an ideal tool for optimizing the imaging and sample preparation conditions.

Paper Details

Date Published: 21 February 2017
PDF: 6 pages
Proc. SPIE 10071, Single Molecule Spectroscopy and Superresolution Imaging X, 100710X (21 February 2017); doi: 10.1117/12.2250116
Show Author Affiliations
Miklós Erdélyi, Univ. of Szeged (Hungary)
József Sinkó, Univ. of Szeged (Hungary)
Tamás Gajdos, Univ. of Szeged (Hungary)
Tibor Novák, Univ. of Szeged (Hungary)


Published in SPIE Proceedings Vol. 10071:
Single Molecule Spectroscopy and Superresolution Imaging X
Jörg Enderlein; Ingo Gregor; Zygmunt Karol Gryczynski; Rainer Erdmann; Felix Koberling, Editor(s)

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