Fig. 2
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 μm3; recording interval: 30 min; scale bars: Fusion, Migration—50 μm, Luminal dynamics—100 μm, 50 μm (inset)