Multi-omics studies, represented by connectomes and spatial transcriptomes, have entered the era of single-cell resolution, necessitating a reference brain atlas with spatial localization capability at the single-cell level1–4. However, such atlases are unavailable5. Here we present a whole mouse brain dataset of Nissl-based cytoarchitecture with isotropic 1-μm resolution, achieved through continuous micro-optical sectioning tomography. By integrating multi-modal images, we constructed a three-dimensional reference atlas of the mouse brain, providing the three-dimensional topographies of 916 structures and enabling arbitrary-angle slice image generation at 1-μm resolution. We developed an informatics-based platform for visualizing and sharing of the atlas images, offering services such as brain slice registration, neuronal circuit mapping and intelligent stereotaxic surgery planning. This atlas is interoperable with widely used stereotaxic atlases, supporting cross-atlas navigation of corresponding coronal planes in two dimensions and spatial mapping across atlas spaces in three dimensions. By facilitating the data analysis and visualization for large brain mapping projects, our atlas promises to be a versatile brainsmatics tool for studying the whole brain at single-cell level.
