Fei Chen, Paul W. Tillberg, and Edward S. Boyden describe a novel technique called expansion microscopy (ExM) that enables the physical magnification of biological specimens. By synthesizing a swellable polymer network within a specimen, the specimen can be physically expanded, allowing for the isotropic separation and optical resolution of labels spaced beyond the optical diffraction limit. This process allows for super-resolution imaging with conventional microscopes, achieving effective lateral resolutions of ~70 nm. The authors demonstrate ExM in both cultured cells and brain tissue, showing that it can resolve individual microtubules and synapses with high precision. They also apply ExM to perform three-color super-resolution imaging of a large volume of mouse hippocampus, revealing nanoscale features such as dendritic spines and synaptic structures. ExM offers a new modality for super-resolution imaging, enabling faster and more detailed analysis of biological samples.Fei Chen, Paul W. Tillberg, and Edward S. Boyden describe a novel technique called expansion microscopy (ExM) that enables the physical magnification of biological specimens. By synthesizing a swellable polymer network within a specimen, the specimen can be physically expanded, allowing for the isotropic separation and optical resolution of labels spaced beyond the optical diffraction limit. This process allows for super-resolution imaging with conventional microscopes, achieving effective lateral resolutions of ~70 nm. The authors demonstrate ExM in both cultured cells and brain tissue, showing that it can resolve individual microtubules and synapses with high precision. They also apply ExM to perform three-color super-resolution imaging of a large volume of mouse hippocampus, revealing nanoscale features such as dendritic spines and synaptic structures. ExM offers a new modality for super-resolution imaging, enabling faster and more detailed analysis of biological samples.