title: Correcting spherical aberrations in a biospecimen using a transmissive liquid crystal device in two-photon excitation laser scanning microscopy.
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.
authors.affiliation: Hokkaido University, Research Institute for Electronic Science, N20W10, Kita-Ward, Sapporo, Hokkaido 001-0020, JapanbHokkaido University, Graduate School of Information Science and Technology, N14W9, Kita-Ward, Sapporo, Hokkaido 060-0814, JapancCitizen Ho.
authors.forename: Ayano
authors.initials: A
authors.lastname: Tanabe
authors.affiliation: Hokkaido University, Research Institute for Electronic Science, N20W10, Kita-Ward, Sapporo, Hokkaido 001-0020, JapanbHokkaido University, Graduate School of Information Science and Technology, N14W9, Kita-Ward, Sapporo, Hokkaido 060-0814, Japan.
authors.forename: Terumasa
authors.initials: T
authors.lastname: Hibi
authors.affiliation: Hokkaido University, Research Institute for Electronic Science, N20W10, Kita-Ward, Sapporo, Hokkaido 001-0020, JapanbHokkaido University, Graduate School of Information Science and Technology, N14W9, Kita-Ward, Sapporo, Hokkaido 060-0814, Japan.
authors.forename: Sari
authors.initials: S
authors.lastname: Ipponjima
authors.affiliation: Citizen Holdings Co. Ltd., 840, Shimotomi, Tokorozawa, Saitama 359-8511, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Matsumoto
authors.affiliation: Citizen Holdings Co. Ltd., 840, Shimotomi, Tokorozawa, Saitama 359-8511, Japan.
authors.forename: Masafumi
authors.initials: M
authors.lastname: Yokoyama
authors.affiliation: Citizen Holdings Co. Ltd., 840, Shimotomi, Tokorozawa, Saitama 359-8511, Japan.
authors.forename: Makoto
authors.initials: M
authors.lastname: Kurihara
authors.affiliation: Citizen Holdings Co. Ltd., 840, Shimotomi, Tokorozawa, Saitama 359-8511, Japan.
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Hashimoto
authors.affiliation: Hokkaido University, Research Institute for Electronic Science, N20W10, Kita-Ward, Sapporo, Hokkaido 001-0020, JapanbHokkaido University, Graduate School of Information Science and Technology, N14W9, Kita-Ward, Sapporo, Hokkaido 060-0814, Japan.
authors.forename: Tomomi
authors.initials: T
authors.lastname: Nemoto
mesh_terms: D016477:Artifacts; D004867:Equipment Design; D019544:Equipment Failure Analysis; D007089:Image Enhancement / Q000295:instrumentation; D007909:Lenses; D050866:Liquid Crystals; D018613:Microscopy, Confocal / Q000295:instrumentation*; D036641:Microscopy, Fluorescence, Multiphoton / Q000295:instrumentation*; D012031:Refractometry / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity