A1 Vertaisarvioitu alkuperäisartikkeli tieteellisessä lehdessä
A software tool for tomographic axial superresolution in STED microscopy
Tekijät: Koho S, Deguchi T, Hanninen PE
Kustantaja: WILEY-BLACKWELL
Julkaisuvuosi: 2015
Journal: Journal of Microscopy
Tietokannassa oleva lehden nimi: JOURNAL OF MICROSCOPY
Lehden akronyymi: J MICROSC-OXFORD
Vuosikerta: 260
Numero: 2
Aloitussivu: 208
Lopetussivu: 218
Sivujen määrä: 11
ISSN: 0022-2720
DOI: https://doi.org/10.1111/jmi.12287
Lay Description STimulated Emission Depletion (STED) microscopy provides intrinsic subdiffraction resolution in far-field fluorescence microscopy. However, the standard lateral donut' STED depletion beam only improves the resolution in the lateral direction, while the axial dimension has the same resolution as a regular confocal microscope. In a typical STED system lateral resolution, the minimum distance between two adjacent image features can be as low as few tens of nanometres, but the axial resolution is limited to 500600 nm - a mismatch of more than an order of magnitude. We propose a new method to deal with this problem, on such axially challenged STED systems, which does not require any modifications of the microscope optics. It is based on tomographic reconstruction of multiple three-dimensional axial projections. Our own STED< method, based on the use of a micromirror placed on top of a standard microscopic sample, is used to record a three-dimensional (3D) projection at an oblique angle in relation to the main optical axis. Combining the STED< projection with the regular STED image into a single view by tomographic reconstruction, is shown to result in a tomogram with significantly improved apparent axial resolution. Our automatic tomographic reconstruction method is demonstrated to work with real biological STED superresolution images, including a data set with a very limited SNR; the reconstruction software (SuperTomo) and its source code will be released under BSD open-source license.