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Resolution in 3D in multifocal plane microscopy

Resolution in 3D in multifocal plane microscopy
Resolution in 3D in multifocal plane microscopy

Using single molecule microscopy, biological interactions can be imaged and studied at the level of individual biomolecules. When characterizing an imaged biological interaction, the distance separating the two participating biomolecules can provide valuable information. Therefore, the resolvability of an imaging setup is of practical significance in the analysis of the acquired image data. Importantly, the resolvability of the imaging setup needs evaluation in the 3D context, since in general biomolecules reside in 3D space within the cellular environment. We recently introduced an information-theoretic 2D resolution measure which shows that the resolution limit due to Rayleigh's criterion can be overcome. This new result predicts that the resolution of optical microscopes is not limited, but rather can be improved with increased photon counts detected from the single molecules. The 2D result was subsequently extended to the 3D context, and the proposed information-theoretic 3D resolution measure can readily be used to determine the resolvability of a conventional single focal plane imaging setup. Here, we consider the 3D resolution measure for a multifocal plane microscope setup, an imaging system which allows the concurrent imaging of multiple focal planes within a specimen. The technique is useful in applications such as the tracking of subcellular objects in 3D. By comparing their 3D resolution measures, we find a two-plane setup to outperform a comparable conventional single-plane setup in resolvability over a range of axial locations for the single molecule pair. Moreover, we investigate and compare the impact of noise on the resolvability of the two setups.

3D resolution measure, Cramer-rao inequality, Fisher information matrix, Multifocal plane microscopy
SPIE
Chao, Jerry
550e20b0-8365-42e3-a6fc-1048eb8c2e47
Ram, Sripad
559bd560-3817-4e53-8c7a-2f08e4518412
Abraham, Anish V.
4f71ee5b-b0b1-4e0e-8f64-5cea9ae8f0e0
Ward, E. Sally
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ober, Raimund J.
31f4d47f-fb49-44f5-8ff6-87fc4aff3d36
Chao, Jerry
550e20b0-8365-42e3-a6fc-1048eb8c2e47
Ram, Sripad
559bd560-3817-4e53-8c7a-2f08e4518412
Abraham, Anish V.
4f71ee5b-b0b1-4e0e-8f64-5cea9ae8f0e0
Ward, E. Sally
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ober, Raimund J.
31f4d47f-fb49-44f5-8ff6-87fc4aff3d36

Chao, Jerry, Ram, Sripad, Abraham, Anish V., Ward, E. Sally and Ober, Raimund J. (2008) Resolution in 3D in multifocal plane microscopy. In Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XV. vol. 6861, SPIE.. (doi:10.1117/12.763084).

Record type: Conference or Workshop Item (Paper)

Abstract

Using single molecule microscopy, biological interactions can be imaged and studied at the level of individual biomolecules. When characterizing an imaged biological interaction, the distance separating the two participating biomolecules can provide valuable information. Therefore, the resolvability of an imaging setup is of practical significance in the analysis of the acquired image data. Importantly, the resolvability of the imaging setup needs evaluation in the 3D context, since in general biomolecules reside in 3D space within the cellular environment. We recently introduced an information-theoretic 2D resolution measure which shows that the resolution limit due to Rayleigh's criterion can be overcome. This new result predicts that the resolution of optical microscopes is not limited, but rather can be improved with increased photon counts detected from the single molecules. The 2D result was subsequently extended to the 3D context, and the proposed information-theoretic 3D resolution measure can readily be used to determine the resolvability of a conventional single focal plane imaging setup. Here, we consider the 3D resolution measure for a multifocal plane microscope setup, an imaging system which allows the concurrent imaging of multiple focal planes within a specimen. The technique is useful in applications such as the tracking of subcellular objects in 3D. By comparing their 3D resolution measures, we find a two-plane setup to outperform a comparable conventional single-plane setup in resolvability over a range of axial locations for the single molecule pair. Moreover, we investigate and compare the impact of noise on the resolvability of the two setups.

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More information

e-pub ahead of print date: 12 February 2008
Published date: 2008
Venue - Dates: Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XV, San Jose, CA, United States, 2008-01-21 - 2008-01-24
Keywords: 3D resolution measure, Cramer-rao inequality, Fisher information matrix, Multifocal plane microscopy

Identifiers

Local EPrints ID: 423596
URI: https://eprints.soton.ac.uk/id/eprint/423596
PURE UUID: bd5e801e-bb1a-45de-9edb-368af2554b0b
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: 29 Oct 2019 01:23

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Contributors

Author: Jerry Chao
Author: Sripad Ram
Author: Anish V. Abraham
Author: E. Sally Ward ORCID iD
Author: Raimund J. Ober ORCID iD

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