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Proceedings Paper

Open-loop wavefront sensing scheme for specimen aberrations correction in two-photon excited fluorescence microscopy
Author(s): Rodrigo Aviles-Espinosa; Jordi Andilla; Rafael Porcar-Guezenec; Xavier Levecq; David Artigas; Pablo Loza-Alvarez
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Paper Abstract

The recent linkage between adaptive optics, a technique borrowed from astronomy and various imaging devices, has enabled to push forward their imaging capabilities by improving its contrast and resolution. A specific case is nonlinear microscopy (NLM) that, although it brings several inherent advantages (compared to linear fluorescence techniques) due to its nonlinear dependence on the excitation beam, its enhanced capabilities can be limited by the sample inhomogeneous structure. In this work, we demonstrate how these imaging capabilities can be enhanced by, employing adaptive optics in a two step correction process. Firstly, a closed-loop methodology aided by Shack-Hartman Wavefront sensing scheme is implemented for compensating the aberrations produced by the laser and the optical elements before the high numerical aperture microscope objective, resulting in a one-time calibration process. Then the residual aberrations are produced by the microscope objective and the sample. These are measured in a similar way as it is done in astronomy (employing a laser guide-star), using the two-photon excited fluorescence. The properties of this incoherent emission produced inside a test sample are compared to a genetically modified Caenorhabditis. elegans nematode expressing GFP showing that the emission of this protein (at 810nm) can be sensed efficiently with our WFS by modifying the exposure time. Therefore the recorded wavefront will capture the sample aberrations which are used to shape a deformable mirror in an open-loop configuration. This correction principle is demonstrated in a test sample by correcting aberrations in a "single-shot" resulting in a reduced sample exposure.

Paper Details

Date Published: 8 June 2011
PDF: 6 pages
Proc. SPIE 8086, Advanced Microscopy Techniques II, 808603 (8 June 2011); doi: 10.1117/12.889397
Show Author Affiliations
Rodrigo Aviles-Espinosa, ICFO-The Institute of Photonic Sciences (Spain)
Jordi Andilla, Imagine Optic (France)
Rafael Porcar-Guezenec, Imagine Optic (France)
Xavier Levecq, Imagine Optic (France)
David Artigas, ICFO-The Institute of Photonic Sciences (Spain)
Univ. Politècnica de Catalunya (Spain)
Pablo Loza-Alvarez, ICFO-The Institute of Photonic Sciences (Spain)

Published in SPIE Proceedings Vol. 8086:
Advanced Microscopy Techniques II
Peter T. C. So; Emmanuel Beaurepaire, Editor(s)

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