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Proceedings Paper

Numerical spherical aberration correction method using spatial light modulator under deep-part fluorescence observation
Author(s): Yu Takiguchi; Hisayoshi Takamoto; Masamitsu Kanada; Takashi Inoue; Naoya Matsumoto; Susumu Terakawa
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Paper Abstract

We have developed a confocal fluorescence laser scanning microscopy (CFLSM) incorporating a liquid crystal on silicon spatial light modulator (LCOS-SLM). To achieve high-resolution and high-contrast imaging for deeper part of the tissue with CFLSM, high numerical aperture objective lenses are required to tightly focus excitation light to meet Rayleigh limit(criterion) for the specimens. However, mismatch of refractive index at the boundary of interfacing materials, such as atmosphere, glass cover, and biological tissues, causes spherical aberration. Recently, we proposed a numerical method for correcting spherical aberration. In this method a pre-distorted wavefront pattern for aberration correction is calculated by ray tracing from a hypothetical focal point inside a specimen to the pupil plane. The resulting microscope can correct such spherical aberration. We observed 6.0μm fluorescent micro-beads dispersed three-dimensionally in agarose gel to confirm effectiveness of aberration correction. We reconstructed a three-dimensional image by taking 20 images by changing the depth with 1 μm interval and stacking them. It was apparent that the longitudinal/depth resolution was improved and that the intensity of fluorescence image was increased with aberration correction. While this method is applicable to other laser scanning microscopes, it has potential to enhance the signals for various super-resolution microscopic techniques, such as stimulated- emission-depletion (STED) fluorescence microscopy.

Paper Details

Date Published: 12 March 2014
PDF: 7 pages
Proc. SPIE 8949, Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXI, 89490C (12 March 2014); doi: 10.1117/12.2038394
Show Author Affiliations
Yu Takiguchi, Hamamatsu Photonics K.K. (Japan)
Hisayoshi Takamoto, Hamamatsu Photonics K.K. (Japan)
Masamitsu Kanada, Hamamatsu Univ. School of Medicine (Japan)
Takashi Inoue, Hamamatsu Photonics K.K. (Japan)
Naoya Matsumoto, Hamamatsu Photonics K.K. (Japan)
Susumu Terakawa, Hamamatsu Univ. School of Medicine (Japan)


Published in SPIE Proceedings Vol. 8949:
Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXI
Thomas G. Brown; Carol J. Cogswell; Tony Wilson, Editor(s)

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