title: Fast imaging of live organisms with sculpted light sheets.
abstract: Light-sheet microscopy is an increasingly popular technique in the life sciences due to its fast 3D imaging capability of fluorescent samples with low photo toxicity compared to confocal methods. In this work we present a new, fast, flexible and simple to implement method to optimize the illumination light-sheet to the requirement at hand. A telescope composed of two electrically tuneable lenses enables us to define thickness and position of the light-sheet independently but accurately within milliseconds, and therefore optimize image quality of the features of interest interactively. We demonstrated the practical benefit of this technique by 1) assembling large field of views from tiled single exposure each with individually optimized illumination settings; 2) sculpting the light-sheet to trace complex sample shapes within single exposures. This technique proved compatible with confocal line scanning detection, further improving image contrast and resolution. Finally, we determined the effect of light-sheet optimization in the context of scattering tissue, devising procedures for balancing image quality, field of view and acquisition speed.
authors.affiliation: Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge, CB2 3RA, UK.|Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge, CB2 3DY, UK.
authors.forename: Aleksander K
authors.initials: AK
authors.lastname: Chmielewski
authors.affiliation: Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge, CB2 3RA, UK.
authors.forename: Anders
authors.initials: A
authors.lastname: Kyrsting
authors.affiliation: Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge, CB2 3RA, UK.
authors.forename: Pierre
authors.initials: P
authors.lastname: Mahou
authors.affiliation: Department of Zoology, University of Cambridge, Downing Street Cambridge, CB2 3EJ, UK.
authors.forename: Matthew T
authors.initials: MT
authors.lastname: Wayland
authors.affiliation: Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge, CB2 3DY, UK.
authors.forename: Leila
authors.initials: L
authors.lastname: Muresan
authors.affiliation: Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge, CB2 3DY, UK.|Center for Organismal Studies, University of Heidelberg, Im Neuenheimer Feld 329, 69120 Heidelberg, Germany.
authors.forename: Jan Felix
authors.initials: JF
authors.lastname: Evers
authors.affiliation: Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge, CB2 3RA, UK.
authors.forename: Clemens F
authors.initials: CF
authors.lastname: Kaminski
mesh_terms: D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism / Q000473:pathology; D004625:Embryo, Nonmammalian / Q000378:metabolism / Q000473:pathology; D007091:Image Processing, Computer-Assisted; D021621:Imaging, Three-Dimensional; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D015027:Zebrafish