ArticleNature protocols2018
Combined expansion microscopy with structured illumination microscopy for analyzing protein complexes.
Article in Nature protocols, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 35 papers.
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Who cites it
35 citing papers in PubMed.
- AI-empowered super-resolution microscopy: a revolution in nanoscale cellular imaging.Nature methods · 2026Review
- ONE microscopy.Nature protocols · 2026Review
- Structure and function of the synaptonemal complex.The Journal of cell biology · 2026Review
- Review of expansion microscopy combined with advanced imaging modalities.Journal of microscopy · 2026Review
- Visualizing Drosophila centrioles by expansion microscopy.Journal of cell science · 2026Article
- Expansion and fluctuations-enhanced microscopy for nanoscale molecular profiling of cells and tissues.Nature protocols · 2026Review
- Deep3DSIM: Super-resolution imaging of thick tissue using 3D structured illumination with adaptive optics.eLife · 2025Article
- Adhesive interactions within microbial consortia can be differentiated at the single-cell level through expansion microscopy.Proceedings of the National Academy of Sciences of the United States of America · 2024Article
- Ultrastructure expansion microscopy (U-ExM) of mouse and human kidneys for analysis of subcellular structures.Cytoskeleton (Hoboken, N.J.) · 2024Article
- Stimuli-Responsive Phosphate Hydrogel: A Study on Swelling Behavior, Mechanical Properties, and Application in Expansion Microscopy.ACS omega · 2024Article
- Adhesive interactions within microbial consortia can be differentiated at the single-cell level through expansion microscopy.bioRxiv : the preprint server for biology · 2024Article
- Expansion microscopy in honeybee brains for high-resolution neuroanatomical analyses in social insects.Cell and tissue research · 2023Article
- Expansion microscopy: A chemical approach for super-resolution microscopy.Current opinion in structural biology · 2023Review
- Superresolution structured illumination microscopy reconstruction algorithms: a review.Light, science & applications · 2023Review
- Far-field super-resolution chemical microscopy.Light, science & applications · 2023Review
- Expansion Microscopy of Mouse Photoreceptor Cilia.Advances in experimental medicine and biology · 2023Article
- Expansion microscopy with carboxylic trifunctional linkers.Acta biochimica et biophysica Sinica · 2022Article
- Nanoscale fluorescence imaging of biological ultrastructure via molecular anchoring and physical expansion.Nano convergence · 2022Review
- Simple methods for quantifying super-resolved cortical actin.Scientific reports · 2022Article
- Technological advances in super-resolution microscopy to study cellular processes.Molecular cell · 2022Review
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Authors and funding
8 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Biologists have long been fascinated with the organization and function of intricate protein complexes. Therefore, techniques for precisely imaging protein complexes and the location of proteins within these complexes are critically important and often require multidisciplinary collaboration. A challenge in these explorations is the limited resolution of conventional light microscopy. However, a new microscopic technique has circumvented this resolution limit by making the biological sample larger, thus allowing for super-resolution of the enlarged structure. This 'expansion' is accomplished by embedding the sample in a hydrogel that, when exposed to water, uniformly expands. Here, we present a protocol that transforms thick expansion microscopy (ExM) hydrogels into sections that are physically expanded four times, creating samples that are compatible with the super-resolution technique structured illumination microscopy (SIM). This super-resolution ExM method (ExM-SIM) allows the analysis of the three-dimensional (3D) organization of multiprotein complexes at ~30-nm lateral (xy) resolution. This protocol details the steps necessary for analysis of protein localization using ExM-SIM, including antibody labeling, hydrogel preparation, protease digestion, post-digestion antibody labeling, hydrogel embedding with tissue-freezing medium (TFM), cryosectioning, expansion, image alignment, and particle averaging. We have used this approach for 3D mapping of in situ protein localization in the Drosophila synaptonemal complex (SC), but it can be readily adapted to study thick tissues such as brain and organs in various model systems. This procedure can be completed in 5 d.
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