Fig. 3.

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 [22]. 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 μm³; recording interval: 30 min; scale bar: 100 μm