The dataset contains raw imaging data from the work: "Long-term live imaging and multiscale analysis identify heterogeneity and core principles of epithelial organoid morphogenesis" The dataset is organized as the following: the "FigureX_" or SupplementaryFigure_X" suffix in the filename refers to the figure in the paper in which the raw data is analyzed and/or visualized. The data is "raw", i.e. not processed. However, in many cases, maximum projections of the original 3D image stacks have been uploaded due to size limitations. The total size of the image stacks approaches 0.5TB. To access the full 3D image stacks please contact the corresponding author (Francesco Pampaloni, fpampalo@bio.uni-frankfurt.de). Authors Lotta Hof1*, Till Moreth1*, Michael Koch1, Tim Liebisch2, Marina Kurtz3, Julia Tarnick4, Susanna M. Lissek5, Monique M.A. Verstegen6, Luc J.W. van der Laan6, Meritxell Huch7, Franziska Matthäus2, Ernst H.K. Stelzer1, Francesco Pampaloni1§ 1Physical Biology Group, Buchmann Institute for Molecular Life Sciences (BMLS), Goethe-Universität Frankfurt am Main, Frankfurt am Main, Germany 2Faculty of Biological Sciences, Goethe-Universität Frankfurt am Main, Frankfurt am Main, Germany 3Department of Physics, Goethe-Universität Frankfurt am Main, Frankfurt am Main, Germany 4Deanery of Biomedical Science, University of Edinburgh, Edinburgh, United Kingdom 5Experimental Medicine and Therapy Research, University of Regensburg, Regensburg, Germany 6Department of Surgery, Erasmus MC – University Medical Center, Rotterdam, The Netherlands 7The Wellcome Trust/CRUK Gurdon Institute, University of Cambridge, Cambridge, United Kingdom. Present address: Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany *contributed equally §corresponding author: fpampalo@bio.uni-frankfurt.de Abstract Background Organoids are morphologically heterogeneous three-dimensional cell culture systems and serve as an ideal model for understanding the principles of collective cell behaviour in mammalian organs during development, homeostasis, regeneration and pathogenesis. To investigate the underlying cell organisation principles of organoids, we imaged hundreds of pancreas and cholangio carcinoma organoids in parallel using light sheet and bright field microscopy for up to seven days. Results We quantified organoid behaviour at single-cell (microscale), individual-organoid (mesoscale), and entire-culture (macroscale) levels. At single-cell resolution, we monitored formation, monolayer polarisation and degeneration, and identified diverse behaviours, including lumen expansion and decline (size oscillation), migration, rotation and multi-organoid fusion. Detailed individual organoid quantifications lead to a mechanical 3D agent-based model. A derived scaling law and simulations support the hypotheses that size oscillations depend on organoid properties and cell division dynamics, which is confirmed by bright field microscopy analysis of entire cultures. Conclusion Our multiscale analysis provides a systematic picture of the diversity of cell organisation in organoids by identifying and quantifying the core regulatory principles of organoid morphogenesis.