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5 August 2015 Correcting spherical aberrations in a biospecimen using a transmissive liquid crystal device in two-photon excitation laser scanning microscopy
Ayano Tanabe, Terumasa Hibi, Sari Ipponjima, Kenji Matsumoto, Masafumi Yokoyama, Makoto Kurihara, Nobuyuki Hashimoto, Tomomi Nemoto
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Journal of Biomedical Optics, Vol. 20, Issue 10, 101204 (August 2015). https://doi.org/10.1117/1.JBO.20.10.101204
Abstract
Two-photon excitation laser scanning microscopy has enabled the visualization of deep regions in a biospecimen. However, refractive-index mismatches in the optical path cause spherical aberrations that degrade spatial resolution and the fluorescence signal, especially during observation at deeper regions. Recently, we developed transmissive liquid-crystal devices for correcting spherical aberration without changing the basic design of the optical path in a conventional laser scanning microscope. In this study, the device was inserted in front of the objective lens and supplied with the appropriate voltage according to the observation depth. First, we evaluated the device by observing fluorescent beads in single- and two-photon excitation laser scanning microscopes. Using a 25× water-immersion objective lens with a numerical aperture of 1.1 and a sample with a refractive index of 1.38, the device recovered the spatial resolution and the fluorescence signal degraded within a depth of ±0.6  mm. Finally, we implemented the device for observation of a mouse brain slice in a two-photon excitation laser scanning microscope. An optical clearing reagent with a refractive index of 1.42 rendered the fixed mouse brain transparent. The device improved the spatial resolution and the yellow fluorescent protein signal within a depth of 0–0.54 mm.
CC BY: © The Authors. Published by SPIE under a Creative Commons Attribution 4.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.
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Ayano Tanabe, Terumasa Hibi, Sari Ipponjima, Kenji Matsumoto, Masafumi Yokoyama, Makoto Kurihara, Nobuyuki Hashimoto, and Tomomi Nemoto "Correcting spherical aberrations in a biospecimen using a transmissive liquid crystal device in two-photon excitation laser scanning microscopy," Journal of Biomedical Optics 20(10), 101204 (5 August 2015). https://doi.org/10.1117/1.JBO.20.10.101204
Published: 5 August 2015
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Cited by 23 scholarly publications.
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KEYWORDS
Monochromatic aberrations
Objectives
Luminescence
Spatial resolution
Laser scanners
Refractive index
LCDs
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