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Optics Express

Optics Express

  • Editor: C. Martijn de Sterke
  • Vol. 17, Iss. 8 — Apr. 13, 2009
  • pp: 6829–6848

Super-resolution orientation estimation and localization of fluorescent dipoles using 3-D steerable filters

François Aguet, Stefan Geissbühler, Iwan Märki, Theo Lasser, and Michael Unser  »View Author Affiliations

Optics Express, Vol. 17, Issue 8, pp. 6829-6848 (2009)

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Fluorophores that are fixed during image acquisition produce a diffraction pattern that is characteristic of the orientation of the fluorophore’s underlying dipole. Fluorescence localization microscopy techniques such as PALM and STORM achieve super-resolution by applying Gaussian-based fitting algorithms to in-focus images of individual fluorophores; when applied to fixed dipoles, this can lead to a bias in the range of 5–20 nm. We introduce a method for the joint estimation of position and orientation of dipoles, based on the representation of a physically realistic image formation model as a 3-D steerable filter. Our approach relies on a single, defocused acquisition. We establish theoretical, localization-based resolution limits on estimation accuracy using Cramér-Rao bounds, and experimentally show that estimation accuracies of at least 5 nm for position and of at least 2 degrees for orientation can be achieved. Patterns generated by applying the image formation model to estimated position/orientation pairs closely match experimental observations.

© 2009 Optical Society of America

OCIS Codes
(070.5010) Fourier optics and signal processing : Pattern recognition
(100.6640) Image processing : Superresolution
(180.2520) Microscopy : Fluorescence microscopy

ToC Category:

Original Manuscript: February 25, 2009
Revised Manuscript: April 3, 2009
Manuscript Accepted: April 3, 2009
Published: April 9, 2009

Virtual Issues
Vol. 4, Iss. 6 Virtual Journal for Biomedical Optics

François Aguet, Stefan Geissbühler, Iwan Märki, Theo Lasser, and Michael Unser, "Super-resolution orientation estimation and localization of fluorescent dipoles using 3-D steerable filters," Opt. Express 17, 6829-6848 (2009)

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