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Fluorescent microspheres as point sources: A localization study

Fluorescent microspheres as point sources: A localization study
Fluorescent microspheres as point sources: A localization study

The localization of fluorescent microspheres is often employed for drift correction and image registration in single molecule microscopy, and is commonly carried out by fitting a point spread function to the image of the given microsphere. The mismatch between the point spread function and the image of the microsphere, however, calls into question the suitability of this localization approach. To investigate this issue, we subject both simulated and experimental microsphere image data to a maximum likelihood estimator that localizes a microsphere by fitting an Airy pattern to its image, and assess the suitability of the approach by evaluating the ability of the estimator to recover the true location of the microsphere with the best possible accuracy as determined based on the Cramér-Rao lower bound. Assessing against criteria based on the standard errors of the mean and the variance for an ideal estimator of the microsphere's location, we find that microspheres up to 100 nm in diameter can in general be localized using a fixed width Airy pattern, and that microspheres as large as 1 μm in diameter can in general be localized using a floated width Airy pattern.

1932-6203
Chao, Jerry
550e20b0-8365-42e3-a6fc-1048eb8c2e47
Lee, Taiyoon
763ca47b-a0fd-43e9-9604-ea148726fa6e
Ward, E. Sally
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ober, Raimund J.
31f4d47f-fb49-44f5-8ff6-87fc4aff3d36
Chao, Jerry
550e20b0-8365-42e3-a6fc-1048eb8c2e47
Lee, Taiyoon
763ca47b-a0fd-43e9-9604-ea148726fa6e
Ward, E. Sally
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ober, Raimund J.
31f4d47f-fb49-44f5-8ff6-87fc4aff3d36

Chao, Jerry, Lee, Taiyoon, Ward, E. Sally and Ober, Raimund J. (2015) Fluorescent microspheres as point sources: A localization study. PLoS ONE, 10 (7), [e0134112]. (doi:10.1371/journal.pone.0134112).

Record type: Article

Abstract

The localization of fluorescent microspheres is often employed for drift correction and image registration in single molecule microscopy, and is commonly carried out by fitting a point spread function to the image of the given microsphere. The mismatch between the point spread function and the image of the microsphere, however, calls into question the suitability of this localization approach. To investigate this issue, we subject both simulated and experimental microsphere image data to a maximum likelihood estimator that localizes a microsphere by fitting an Airy pattern to its image, and assess the suitability of the approach by evaluating the ability of the estimator to recover the true location of the microsphere with the best possible accuracy as determined based on the Cramér-Rao lower bound. Assessing against criteria based on the standard errors of the mean and the variance for an ideal estimator of the microsphere's location, we find that microspheres up to 100 nm in diameter can in general be localized using a fixed width Airy pattern, and that microspheres as large as 1 μm in diameter can in general be localized using a floated width Airy pattern.

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journal.pone.0134112 - Version of Record
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Accepted/In Press date: 6 July 2015
e-pub ahead of print date: 28 July 2015
Published date: 28 July 2015

Identifiers

Local EPrints ID: 423663
URI: http://eprints.soton.ac.uk/id/eprint/423663
ISSN: 1932-6203
PURE UUID: 36b48a93-622e-4cd1-b347-1cb922d46f95
ORCID for E. Sally Ward: ORCID iD orcid.org/0000-0003-3232-7238
ORCID for Raimund J. Ober: ORCID iD orcid.org/0000-0002-1290-7430

Catalogue record

Date deposited: 27 Sep 2018 16:30
Last modified: 20 Jul 2019 00:22

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Contributors

Author: Jerry Chao
Author: Taiyoon Lee
Author: E. Sally Ward ORCID iD
Author: Raimund J. Ober ORCID iD

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