SynAtlas: A Safe and Versatile Platform for Rapid Organ‐Scale Tissue Clearing and 3D Atlas Mapping

Understanding the detailed three-dimensional organization of cells, molecules, and tissues in their natural state is essential for advancing knowledge of biology, disease, and neural circuits, yet traditional slicing methods are slow, destructive, and prone to artifacts, while existing tissue-clearing techniques often compromise speed, safety, signal preservation, or simplicity. By systematically analyzing the distinct optical barriers that impede tissue transparency, we propose an orthogonal synergy design principle. Guided by this framework, we developed SynAtlas (Synergistic Aqueous Tissue clearing System), a rationally designed clearing method that combines safe, compatible chemicals to simultaneously overcome the main barriers to transparency-lipid scattering, protein hydration heterogeneity, and refractive index mismatch-creating a fast, modular, and low-hazard platform. SynAtlas clears thin sections in minutes and whole organs within two days while excellently preserving fluorescent proteins, nucleic acids, and antibody labels; it enables high-resolution 3D atlases of neurons, vasculature, gene expression patterns, and cell types in mouse organs and archival human brain tissue, supports multi-round labeling with minimal distortion, and allows non-destructive cryo-preservation for long-term storage. This versatile approach makes advanced volumetric imaging more accessible and impactful for research and clinical applications.