Super-Resolution Imaging of Intracellular Molecules Using Expansion Microscopy

Abstract

Expansion Microscopy (ExM) is a super-resolution imaging technique that improves resolution by isotropic physical expansion of biological samples embedded in a swollen hydrogel. As a result, ExM enables nanoscale imaging using conventional fluorescence microscopy. A decade following the inception of ExM, comprehensive spatial analysis of proteins, nucleic acids (RNA and DNA), lipids, glycan chains, and other biomolecules is now applicable to a broad spectrum of cellular and biological contexts. Recent methodological advances, such as increased expansion factors, improved labeling efficiency, and more efficient and cost-effective sample preparation protocols, have accelerated their application and facilitated many scientific discoveries. This review article focuses on the basic principles underlying ExM, identifies practical protocols for its implementation and observation, highlights recent technological advances, and discusses remaining challenges. Ultimately, it aims to provide a comprehensive overview of ExM to encourage its broad application and methodological innovation.