FOCUS-3D: Robust, generalizable volumetric cell segmentation for three-dimensional fluorescence microscopy

Understanding how cells establish spatial organization within tissues is a fundamental question in life sciences. While modern three-dimensional fluorescence microscopy captures large-volume tissue architecture, extracting quantitative cellular insights from complex volumetric datasets remains a major barrier. Here, we introduce FOCUS-3D, a robust, broadly generalizable volumetric cell segmentation framework built on a large, diverse manually annotated cell resource and advanced AI designs. Integrating volumetric representation learning, multi-scale feature extraction, and query-based mask prediction, FOCUS-3D achieves state-of-the-art performance across diverse species, tissues, fluorescent reporters and imaging modalities. During zebrafish (Danio rerio) development, FOCUS-3D uncovers three successive phases of notochord morphogenesis. We disentangle early motility-driven rearrangements from later cell shape remodeling and tissue repacking, and further link these morphological states to spatial and developmental transcriptional programs across independent datasets.