Fig. 1: Light-sheet microscopy provides fast imaging with minimized photo-toxicity and photo-bleaching, enabling diverse applications from imaging developmental processes to imaging of large, cleared tissues.
From: Light-sheets and smart microscopy, an exciting future is dawning
a Traditional light-sheet microscopy such as three-objective Selective Plane Illumination Microscopy (SPIM) relies on an orthogonal arrangement of the illumination (blue; illumination objectives IL1 and IL2) and detection (green; detection objective DO). This ensures that the axial resolution of imaging is mainly governed by the thickness of the light-sheet (blue) enabling imaging across large samples with widefield detection (green) and good optical sectioning. To acquire a 3D volume, the sample is scanned along the detection axis either by moving the sample itself or by scanning the light sheet together with the objective in the detection path. b–d Prime examples of imaging with light-sheets include continuous, long-term imaging of developmental processes in mouse and zebrafish embryos, and imaging of cleared tissue with subcellular resolution. b Katie McDole et al.4 characterized the cellular movements involved in mouse development from early streak (E6.5) to somite stages (E8.5) by imaging a CAGTAG1 expressing mouse embryo with a histone marker (H2B-eGFP) for over 44 h. Scale bar: 100 μm. c Selected projections from multi-view imaging5 (three angles) of the growing embryonic zebrafish vasculature labeled with the fluorescent vascular marker (Tg(kdrl:EGFP), cyan) and the red blood cell marker (Tg(GATA1a:dsRed), magenta), imaged from 20 h post-fertilization (hpf) to 86 hpf. Scale bar: 500 μm d Adam Glaser et al.37 performed large-scale imaging of an expanded slice of kidney of 3.2 cm × 2.1 cm size and 1 mm thickness. High-resolution regions of interest revealed the morphology of glomeruli (Scale bar: 40 μm), vessels (Scale bar: 80 μm) and tubules (Scale bar: 50 μm). The increased resolution due to expansion was further demonstrated with a multi-channel zoom-in of DAPI-counterstained tissue (Scale bars: 100 μm [top] and 20 μm [bottom]). The scale bars thereby indicate the dimensions of the native unexpanded tissue. Panel b was adapted with permission from Katie McDole et al. (2018)4. Panel c adapted from Daetwyler et al. (2019)5. Panel d adapted from Glaser et al. (2019)37.
