Journal: Frontiers in Neuroscience

Article Title: Rapid and efficient optical tissue clearing for volumetric imaging of the intact and injured spinal cord in mice

doi: 10.3389/fnins.2025.1601360

Figure Lengend Snippet: The sciDISCO protocol allows for the 3D imaging of the entire spinal cord. (A) Visual schematic of the sciDISCO protocol involving the delipidation of the spinal cord, immunolabeling, and RI matching in ethyl cinnamate. Created in BioRender ( https://BioRender.com/7kqz2km ). (B) 3D schematics of the spinal cord straightener and agarose mounting chamber. (C) (i) A curved spinal cord after post-fixation and dissection. (ii) The spinal cord in the 3D printed spinal cord straightener and (iii) the straightened spinal cord after delipidation and removal from the straightener. (D) A cleared spinal cord embedded in agarose and RI matched with ethyl cinnamate. (E) Horizontal view of a 3D lightsheet image of a spinal cord including cervical, thoracic, and lumbar segments labeled with NeuN antibody to visualize neuronal cell bodies. (F) Zoomed in sagittal view of the outline thoracic region (yellow) from (E) . (G) Virtual transverse cross section of the thoracic spinal cord. Grids on images are made up of 2.5 mm × 2.5 mm squares.

Article Snippet: After equilibration in ethyl cinnamate for at least 1 day, spinal cord samples were imaged using a Zeiss Z1 Lightsheet microscope with a 5x acquisition objective.

Techniques: Imaging, Immunolabeling, Dissection, Labeling

Journal: Frontiers in Neuroscience

Article Title: Rapid and efficient optical tissue clearing for volumetric imaging of the intact and injured spinal cord in mice

doi: 10.3389/fnins.2025.1601360

Figure Lengend Snippet: sciDISCO is faster than comparable clearing protocols and delivers successful immunolabeling. (A) Timeline schematic detailing the average times for each main step in sciDISCO, iDISCO+, and the PACT clearing protocols. Created in BioRender ( https://BioRender.com/ghn1jij ). (B) sciDISCO delivers similar immunolabeling of NeuN when compared to iDISCO+ and PACT clearing protocols. Representative maximum intensity projections of 3D lightsheet images of the lower cervical and upper thoracic regions are shown. (C) Representative maximum intensity projections of NeuN-labeled thoracic spinal cord regions cleared using either DCM or BTF in the sciDISCO protocol. (D) Quantification of the signal-to-background ratio of samples cleared with sciDISCO using either DCM or BTF. Mean ± SEM. Student’s t -test. (E) Quantification of mean fluorescent intensity of samples incubated in ethyl cinnamate (sciDISCO) vs. dibenzyl ether (iDISCO+) after 1 and 30 days of incubation. Mean ± SEM. ** p < 0.01. Two-way ANOVA with Tukey’s multiple comparisons post-hoc test.

Article Snippet: After equilibration in ethyl cinnamate for at least 1 day, spinal cord samples were imaged using a Zeiss Z1 Lightsheet microscope with a 5x acquisition objective.

Techniques: Immunolabeling, Labeling, Incubation

Journal: Frontiers in Neuroscience

Article Title: Rapid and efficient optical tissue clearing for volumetric imaging of the intact and injured spinal cord in mice

doi: 10.3389/fnins.2025.1601360

Figure Lengend Snippet: sciDISCO allows for the immunolabeling and imaging of the meningeal layers around the spinal cord. (A) Macroscopic image of a mouse spinal cord cleared and RI matched in RIMS (PACT) with the meninges still intact. The PACT clearing process causes the tissue parenchyma to swell and the meninges to constrict around the spinal cord (arrowhead). (B) NeuN and Lyve1 immunostaining of a PACT cleared spinal cord with meninges removed (top) and intact (bottom). (C) Macroscopic image of a mouse spinal cord cleared and RI matched with ethyl cinnamate (sciDISCO) with the meninges still intact. (D) NeuN and Lyve1 immunostaining of a sciDISCO cleared spinal cord with the meninges removed (top) and with the meninges intact (bottom). All images in (B,D) are maximum intensity projections of 3D lightsheet images. Grids on macroscopic images are made up of 2.5 mm × 2.5 mm squares.

Article Snippet: After equilibration in ethyl cinnamate for at least 1 day, spinal cord samples were imaged using a Zeiss Z1 Lightsheet microscope with a 5x acquisition objective.

Techniques: Immunolabeling, Imaging, Immunostaining

Journal: Frontiers in Neuroscience

Article Title: Rapid and efficient optical tissue clearing for volumetric imaging of the intact and injured spinal cord in mice

doi: 10.3389/fnins.2025.1601360

Figure Lengend Snippet: sciDISCO is compatible with transgenic mouse lines for sparse labeling of neuronal populations. (A) Graphical schematic of the stochastic Confetti fluorescent reporter expression upon crossing with Chx10-Cre mice. Created in BioRender ( https://BioRender.com/5cwn62g ). (B) Lightsheet imaging of RFP + V2a interneurons in the thoracic spinal cord following clearing with sciDISCO. (C) Graphical schematic of the MORF3 sparse labeling system upon crossing with Chx10-Cre mice. Created in BioRender ( https://BioRender.com/fh674nv ). (D) Lightsheet images of very sparse labeling of V2a interneurons in the thoracic region of the spinal cord following sciDISCO clearing. Both images in (B,D) are partial maximum intensity projections of the intermediate laminae of the spinal cord from the 3D lightsheet images.

Article Snippet: After equilibration in ethyl cinnamate for at least 1 day, spinal cord samples were imaged using a Zeiss Z1 Lightsheet microscope with a 5x acquisition objective.

Techniques: Transgenic Assay, Labeling, Expressing, Imaging

Journal: BMC Biology

Article Title: Long-term live imaging and multiscale analysis identify heterogeneity and core principles of epithelial organoid morphogenesis

doi: 10.1186/s12915-021-00958-w

Figure Lengend Snippet: Time-resolved live LSFM recordings for detailed qualitative inspections of dynamic morphological processes in organoid development. hCCAOs and mPOs were seeded into Z1-FEP-cuvettes for long-term live observations. They expressed the nuclei marker H2B-eGFP (magenta) or Rosa26-nTnG (grey) and the F-actin cytoskeletal marker LifeAct-mCherry (green). About 120 organoids were recorded in image stacks up to 900 z -planes deep for at most 7 days. The figure shows excerpts of maximum intensity z -projections. Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 488 nm, 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bars: 50 μm, 25 μm (inset)

Article Snippet: Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 488 nm, 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bars: Fusion, Migration—50 μm, Luminal dynamics—100 μm, 50 μm (inset) Additional file 17: Video 3.

Techniques: Marker, Microscopy, Blocking Assay

Journal: BMC Biology

Article Title: Long-term live imaging and multiscale analysis identify heterogeneity and core principles of epithelial organoid morphogenesis

doi: 10.1186/s12915-021-00958-w

Figure Lengend Snippet: High-quality live LSFM image data provide an excellent basis for volume rendering and detailed feature tracking. It can be used for the quantitative description of cellular dynamics in organoid development. 3D renderings offer detailed views into processes such as organoid fusion and elucidate the spatial context in observed luminal dynamics. 3D cell tracking reveals the complex rotation of the epithelial cell monolayer. hCCAOs (seeded and maintained in Z1-FEP-cuvettes) expressed the nuclei marker H2B-eGFP (magenta) and the F-actin cytoskeletal marker LifeAct-mCherry (green). The figure shows segmented and tracked cell nuclei (Rotation; centroids—red; tracks—rainbow), excerpts of maximum intensity z -projections and 3D renderings of corresponding data sets. Segmentation, tracking, and 3D rendering were performed with Arivis Vision4D. Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 488 nm, 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bars: Fusion, Migration—50 μm, Luminal dynamics—100 μm, 50 μm (inset)

Article Snippet: Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 488 nm, 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bars: Fusion, Migration—50 μm, Luminal dynamics—100 μm, 50 μm (inset) Additional file 17: Video 3.

Techniques: Cell Tracking Assay, Marker, Microscopy, Blocking Assay, Migration

Journal: BMC Biology

Article Title: Long-term live imaging and multiscale analysis identify heterogeneity and core principles of epithelial organoid morphogenesis

doi: 10.1186/s12915-021-00958-w

Figure Lengend Snippet: Long-term single-cell analysis of mPOs reveals heterogeneity of proliferation potentials. mPOs (seeded and maintained in one Z1-FEP-cuvettes) expressed the nuclei marker Rosa26-nTnG (grey). Organoids were imaged for 6 days and analysed with our previously published nuclei segmentation pipeline . a Three representative organoids are shown directly after seeding (0 h, upper row) and after 6 days (144 h, lower row). Every row shows one view of the same Z1-FEP-cuvette. Hence, all displayed organoids were grown simultaneously within one FEP-cuvette. The close-ups display the segmentation of the organoid at the corresponding time point. Different colours refer to individual cell nuclei. The coloured frames indicate organoids with different proliferation rates—green/high, blue/low, and red/medium. b From top to bottom, corresponding evaluations of volume, surface area, and neighbourhood relationships (DCG: Delaunay cell graph; PCG: proximity cell graph). Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bar: 100 μm

Article Snippet: Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 488 nm, 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bars: Fusion, Migration—50 μm, Luminal dynamics—100 μm, 50 μm (inset) Additional file 17: Video 3.

Techniques: Single-cell Analysis, Marker, Microscopy, Blocking Assay

Journal: BMC Biology

Article Title: Long-term live imaging and multiscale analysis identify heterogeneity and core principles of epithelial organoid morphogenesis

doi: 10.1186/s12915-021-00958-w

Figure Lengend Snippet: Volume analysis of three representative mPOs reveals different oscillation frequencies . MPOs (seeded and maintained in one Z1-FEP-cuvettes) expressed the nuclei marker Rosa26-nTnG (grey). Organoids were imaged for 6 days and three representative organoids were analysed with our previously published nuclei segmentation pipeline in regard to size oscillation events. A size oscillation lasts between 30 min and 2 h. a Volume over time for each organoid approximated from the cell nuclei segmentation. b Close-up of three (red), two (green) and seven (blue) size oscillation events with more than 5% volume reductions are shown. c Typical images of organoid size oscillation. The upper row shows the nuclei in grey in the raw image, the lower row the segmented cell nuclei. Each colour illustrates a single-cell nucleus. Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bar: 100 μm

Article Snippet: Microscope: Zeiss Lightsheet Z.1; objective lenses: detection: W Plan-Apochromat × 20/1.0, illumination: Zeiss LSFM × 10/0.2; laser lines: 488 nm, 561 nm; filters: laser block filter (LBF) 405/488/561; voxel size: 1.02 × 1.02 × 2.00 μm 3 ; recording interval: 30 min; scale bars: Fusion, Migration—50 μm, Luminal dynamics—100 μm, 50 μm (inset) Additional file 17: Video 3.

Techniques: Marker, Microscopy, Blocking Assay

Journal: Nature Communications

Article Title: Nuclear speed and cycle length co-vary with local density during syncytial blastoderm formation in a cricket

doi: 10.1038/s41467-022-31212-8

Figure Lengend Snippet: a Time points from the embryonic syncytial development of G. bimaculatus , displayed as \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$z$$\end{document} z -projections of 3D stacks. Micrographs are from an H2B-EGFP transgenic G. bimaculatus embryo live-imaged using a lightsheet microscope over eight hours of development at 28.5 °C, capturing nuclear divisions and movements throughout the syncytial embryo. The nuclei (n) arrange into a single layer, after which cellularization occurs and the embryonic rudiment forms. Embryos are oriented laterally with ventral to the bottom and anterior to the left. Anterior is to the left in all subsequent figures. Embryonic stage (ES) and egg stage (EgS) are indicated for each time point . b Nuclei were tracked to produce a 3D + T dataset of nuclear lineages. All nuclear tracks are displayed for an example embryo, with the lineage descended from a single nucleus highlighted. Color scale represents time; n = number of nuclei. Black arrowhead highlights nuclei that move in a highly directed manner toward the anterior pole. c Example time points from a 3D + T dataset of a transgenic G. bimaculatus embryo with nuclei and cytoplasm fluorescently marked (further details in Methods). Left column shows the nucleus channel, and the right column shows the cytoplasm channel, with the energid cytoplasm highlighted in magenta and the nucleus highlighted in cyan. White arrowheads mark two putative yolk granules that remain in place as the nucleus moves past them. d Pairwise correlations between the instantaneous movement vectors of pairs of non-sister nuclei. White line indicates median, box indicates 25 th −75 th percentiles, whiskers show range. Source data are provided as a Source Data file.

Article Snippet: Lightsheet imaging was conducted with a Zeiss Z.1 Lightsheet microscope controlled by Zen Black software (Zeiss, 2014-2018), with the agarose column immersed in a bath of distilled water, temperature controlled at 28.5 °C.

Techniques: Transgenic Assay, Microscopy

Journal: STAR Protocols

Article Title: 4D light sheet imaging, computational reconstruction, and cell tracking in mouse embryos

doi: 10.1016/j.xpro.2024.103515

Figure Lengend Snippet: Software packages used in this protocol

Article Snippet: Regarding microscopy equipment, preparation of embedding medium (#1–2) and microscope setup (#10–19) were written for direct application to Zeiss Lightsheet Z.1 microscope (and Lightsheet 7), given the upside-down chandalier-like 4-axis stage and glass capillary sample embedding.

Techniques: Software, Imaging, Microscopy, Plasmid Preparation

Journal: STAR Protocols

Article Title: 4D light sheet imaging, computational reconstruction, and cell tracking in mouse embryos

doi: 10.1016/j.xpro.2024.103515

Figure Lengend Snippet:

Article Snippet: Regarding microscopy equipment, preparation of embedding medium (#1–2) and microscope setup (#10–19) were written for direct application to Zeiss Lightsheet Z.1 microscope (and Lightsheet 7), given the upside-down chandalier-like 4-axis stage and glass capillary sample embedding.

Techniques: Recombinant, Saline, Cell Culture, Sterility, Software, Imaging, Microscopy, Incubation, Dissection, Fluorescence, Laser Capture Microdissection