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Optical Clearing and Light Sheet Microscopy Imaging of Amphioxus
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SYSNO ASEP 0555020 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve WOS Title Optical Clearing and Light Sheet Microscopy Imaging of Amphioxus Author(s) Macháčová, Simona (UMG-J)
Chmelová, Helena (UMG-J)
Vávrová, Anna (UMG-J)
Kozmik, Zbyněk (UMG-J) RID
Kozmiková, Iryna (UMG-J) RIDNumber of authors 5 Article number 702986 Source Title Frontiers in Cell and Developmental Biology. - : Frontiers Research Foundation - ISSN 2296-634X
Roč. 9, July (2021)Number of pages 12 s. Publication form Online - E Language eng - English Country CH - Switzerland Keywords amphioxus ; light sheet microscopy ; clearing technique ; whole mount immunohistochemistry ; photoreceptor ; acetylated tubulin ; melanopsin Subject RIV EB - Genetics ; Molecular Biology OECD category Cell biology R&D Projects GA20-25377S GA ČR - Czech Science Foundation (CSF) Method of publishing Open access Institutional support UMG-J - RVO:68378050 UT WOS 000682949500001 DOI 10.3389/fcell.2021.702986 Annotation Cephalochordates (amphioxi or lancelets) are representatives of the most basally divergent group of the chordate phylum. Studies of amphioxus development and anatomy hence provide a key insight into vertebrate evolution. More widespread use of amphioxus in the evo-devo field would be greatly facilitated by expanding the methodological toolbox available in this model system. For example, evo-devo research on amphioxus requires deep understanding of animal anatomy. Although conventional confocal microscopy can visualize transparent amphioxus embryos and early larvae, the imaging of later developmental stages is problematic because of the size and opaqueness of the animal. Here, we show that light sheet microscopy combined with tissue clearing methods enables exploration of large amphioxus specimens while keeping the surface and the internal structures intact. We took advantage of the phenomenon of autofluorescence of amphioxus larva to highlight anatomical details. In order to investigate molecular markers at the single-cell level, we performed antibody-based immunodetection of melanopsin and acetylated-alpha-tubulin to label rhabdomeric photoreceptors and the neuronal scaffold. Our approach that combines light sheet microscopy with the clearing protocol, autofluorescence properties of amphioxus, and antibody immunodetection allows visualizing anatomical structures and even individual cells in the 3D space of the entire animal body. Workplace Institute of Molecular Genetics Contact Nikol Škňouřilová, nikol.sknourilova@img.cas.cz, Tel.: 241 063 217 Year of Publishing 2022 Electronic address https://www.frontiersin.org/articles/10.3389/fcell.2021.702986/full
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