Landscape Expansion Microscopy Reveals Interactions between Membrane and Phase-Separated Organelles

Abstract Landscape Expansion Microscopy (land-ExM) is a light microscopy technique that visualizes both lipid and protein ultrastructural context of cells. Achieving this level of detail requires both superresolution and a high signal-to-noise ratio. Although expansion microscopy (ExM) provides superresolution, obtaining high signal-to-noise images of both proteins and lipids remains challenging. Land-ExM overcomes this limitation by using self-retention trifunctional anchors to significantly enhance protein and lipid signals in expanded samples. This improvement enables the accurate visualization of diverse membrane organelles and phase separations, as well as the three-dimensional visualization of their contact sites. As a demonstration, we revealed triple-organellar contact sites among the stress granule, the nuclear tunnel, and the nucleolus. Overall, land-ExM offers a high-contrast superresolution platform that advances our understanding of how cells spatially coordinate interactions between membrane organelles and phase separations. eTOC Summary Zhuang et al. introduce land-ExM, a super-resolution approach that simultaneously maps protein and lipid ultrastructure in cells with high contrast. This method visualizes 3D interactions between membrane-bound organelles and phase-separated condensates, uncovering organelle contact sites such as stress granules at nuclear tunnels adjacent to nucleoli. Competing Interest Statement X.S is a cofounder of Cytoseen. The remaining authors declare no competing interests.