Localization accuracy in single-molecule microscopy
Localization accuracy in single-molecule microscopy
One of the most basic questions in single-molecule microscopy concerns the accuracy with which the location of a single molecule can be determined. Using the Fisher information matrix it is shown that the limit of the localization accuracy for a single molecule is given by λem/2πn a√γAt, where λem, na, γ, A, and t denote the emission wavelength of the single molecule, the numerical aperture of the objective, the efficiency of the optical system, the emission rate of the single molecule and the acquisition time, respectively. Using Monte Carlo simulations it is shown that estimation algorithms can come close to attaining the limit given in the expression. Explicit quantitative results are also provided to show how the limit of the localization accuracy is reduced by factors such as pixelation of the detector and noise sources in the detection system. The results demonstrate what is achievable by single-molecule microscopy and provide guidelines for experimental design.
1185-1200
Ober, Raimund J.
31f4d47f-fb49-44f5-8ff6-87fc4aff3d36
Ram, Sripad
559bd560-3817-4e53-8c7a-2f08e4518412
Ward, E. Sally
b31c0877-8abe-485f-b800-244a9d3cd6cc
February 2004
Ober, Raimund J.
31f4d47f-fb49-44f5-8ff6-87fc4aff3d36
Ram, Sripad
559bd560-3817-4e53-8c7a-2f08e4518412
Ward, E. Sally
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ober, Raimund J., Ram, Sripad and Ward, E. Sally
(2004)
Localization accuracy in single-molecule microscopy.
Biophysical Journal, 86 (2), .
(doi:10.1016/S0006-3495(04)74193-4).
Abstract
One of the most basic questions in single-molecule microscopy concerns the accuracy with which the location of a single molecule can be determined. Using the Fisher information matrix it is shown that the limit of the localization accuracy for a single molecule is given by λem/2πn a√γAt, where λem, na, γ, A, and t denote the emission wavelength of the single molecule, the numerical aperture of the objective, the efficiency of the optical system, the emission rate of the single molecule and the acquisition time, respectively. Using Monte Carlo simulations it is shown that estimation algorithms can come close to attaining the limit given in the expression. Explicit quantitative results are also provided to show how the limit of the localization accuracy is reduced by factors such as pixelation of the detector and noise sources in the detection system. The results demonstrate what is achievable by single-molecule microscopy and provide guidelines for experimental design.
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Accepted/In Press date: 9 October 2003
Published date: February 2004
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Local EPrints ID: 424081
URI: http://eprints.soton.ac.uk/id/eprint/424081
ISSN: 0006-3495
PURE UUID: 10e36e7e-2347-4980-8366-9cd764726069
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Date deposited: 04 Oct 2018 16:30
Last modified: 16 Mar 2024 04:37
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Author:
Sripad Ram
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