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4Pi fluorescence detection and 3D particle localization with a single objective

Coherent detection through two opposing objectives (4Pi configuration) improves the precision of three-dimensional (3D) single-molecule localization substantially along the axial direction, but suffers from instrument complexity and maintenance difficulty. To address these issues, we have realized 4...

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Detalles Bibliográficos
Autores principales: Schnitzbauer, J., McGorty, R., Huang, B.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Optical Society of America 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3796685/
https://www.ncbi.nlm.nih.gov/pubmed/24105517
http://dx.doi.org/10.1364/OE.21.019701
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author Schnitzbauer, J.
McGorty, R.
Huang, B.
author_facet Schnitzbauer, J.
McGorty, R.
Huang, B.
author_sort Schnitzbauer, J.
collection PubMed
description Coherent detection through two opposing objectives (4Pi configuration) improves the precision of three-dimensional (3D) single-molecule localization substantially along the axial direction, but suffers from instrument complexity and maintenance difficulty. To address these issues, we have realized 4Pi fluorescence detection by sandwiching the sample between the objective and a mirror, and create interference of direct incidence and mirror-reflected signal at the camera with a spatial light modulator. Multifocal imaging using this single-objective mirror interference scheme offers improvement in the axial localization similar to the traditional 4Pi method. We have also devised several PSF engineering schemes to enable 3D localization with a single emitter image, offering better axial precision than normal single-objective localization methods such as astigmatic imaging.
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spelling pubmed-37966852014-08-15 4Pi fluorescence detection and 3D particle localization with a single objective Schnitzbauer, J. McGorty, R. Huang, B. Opt Express Research-Article Coherent detection through two opposing objectives (4Pi configuration) improves the precision of three-dimensional (3D) single-molecule localization substantially along the axial direction, but suffers from instrument complexity and maintenance difficulty. To address these issues, we have realized 4Pi fluorescence detection by sandwiching the sample between the objective and a mirror, and create interference of direct incidence and mirror-reflected signal at the camera with a spatial light modulator. Multifocal imaging using this single-objective mirror interference scheme offers improvement in the axial localization similar to the traditional 4Pi method. We have also devised several PSF engineering schemes to enable 3D localization with a single emitter image, offering better axial precision than normal single-objective localization methods such as astigmatic imaging. Optical Society of America 2013-08-14 /pmc/articles/PMC3796685/ /pubmed/24105517 http://dx.doi.org/10.1364/OE.21.019701 Text en ©2013 Optical Society of America http://creativecommons.org/licenses/by-nc-nd/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 Unported License, which permits download and redistribution, provided that the original work is properly cited. This license restricts the article from being modified or used commercially.
spellingShingle Research-Article
Schnitzbauer, J.
McGorty, R.
Huang, B.
4Pi fluorescence detection and 3D particle localization with a single objective
title 4Pi fluorescence detection and 3D particle localization with a single objective
title_full 4Pi fluorescence detection and 3D particle localization with a single objective
title_fullStr 4Pi fluorescence detection and 3D particle localization with a single objective
title_full_unstemmed 4Pi fluorescence detection and 3D particle localization with a single objective
title_short 4Pi fluorescence detection and 3D particle localization with a single objective
title_sort 4pi fluorescence detection and 3d particle localization with a single objective
topic Research-Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3796685/
https://www.ncbi.nlm.nih.gov/pubmed/24105517
http://dx.doi.org/10.1364/OE.21.019701
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