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Optics Express

Optics Express

  • Editor: Andrew M. Weiner
  • Vol. 21, Iss. 17 — Aug. 26, 2013
  • pp: 19701–19708

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

J. Schnitzbauer, R. McGorty, and B. Huang  »View Author Affiliations

Optics Express, Vol. 21, Issue 17, pp. 19701-19708 (2013)

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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.

© 2013 OSA

OCIS Codes
(110.6880) Imaging systems : Three-dimensional image acquisition
(180.2520) Microscopy : Fluorescence microscopy
(180.3170) Microscopy : Interference microscopy
(350.5730) Other areas of optics : Resolution
(110.1080) Imaging systems : Active or adaptive optics

ToC Category:

Original Manuscript: June 21, 2013
Revised Manuscript: August 6, 2013
Manuscript Accepted: August 7, 2013
Published: August 14, 2013

Virtual Issues
Vol. 8, Iss. 9 Virtual Journal for Biomedical Optics

J. Schnitzbauer, R. McGorty, and B. Huang, "4Pi fluorescence detection and 3D particle localization with a single objective," Opt. Express 21, 19701-19708 (2013)

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