ArticleNature protocols2019
A practical guide to optimization in X10 expansion microscopy.
Article in Nature protocols, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 86 papers.
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Who cites it
86 citing papers in PubMed.
- Multi-immersion oblique plane microscope (miOPM): a reconfigurable platform for high-resolution light-sheet fluorescence microscopy.Nature methods · 2026Article
- Years-long stability of X10 expansion microscopy hydrogels.Journal of biological engineering · 2026Article
- Tissue expansion mass spectrometry imaging (TEMI) for high-spatial-resolution multiomics molecular mapping.Nature protocols · 2026Review
- ONE microscopy.Nature protocols · 2026Review
- Hydrogel Chemistry in Expansion Microscopy: From Polyacrylamide to Next-Generation Swellable Polymers.Chembiochem : a European journal of chemical biology · 2026Review
- Single-Step Expansion SMLM Enables Molecular-Resolution Imaging in Cells and Isolated Proteins.Chemical & biomedical imaging · 2026Article
- A uniform tissue-clearing framework and mesoSPIM-ultra enable cm-scale single-neuron tracing.bioRxiv : the preprint server for biology · 2026Article
- Article
- Filter-aided expansion proteomics for the spatial analysis of single cells and organelles in FFPE tissue samples.Nature protocols · 2026Review
- Article
- Review of expansion microscopy combined with advanced imaging modalities.Journal of microscopy · 2026Review
- Seq-Scope-eXpanded: spatial omics beyond optical resolution.Nature communications · 2026Article
- MISO regulates mitochondrial dynamics and mtDNA homeostasis by establishing membrane subdomains.Nature cell biology · 2026Article
- Principles ofbioRxiv : the preprint server for biology · 2026Article
- Visualising viral interactions and mechanisms at the nanoscale with expansion microscopy.Npj viruses · 2026Review
- Optical Approaches to Dissect the Structure and Dynamics of the Synapse at Nanoscale Resolution.Advances in neurobiology · 2026Review
- Expansion and fluctuations-enhanced microscopy for nanoscale molecular profiling of cells and tissues.Nature protocols · 2026Review
- High Precision Antibody-Free Microtubule Labeling for Expansion Microscopy.Bio-protocol · 2025Article
- Article
- One-step nanoscale expansion microscopy reveals individual protein shapes.Nature biotechnology · 2025Article
26 more citing papers are in PubMed but not listed here.
Corrections and comments
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Authors and funding
4 authors.
Funding
Abstract
Expansion microscopy is a relatively new approach to super-resolution imaging that uses expandable hydrogels to isotropically increase the physical distance between fluorophores in biological samples such as cell cultures or tissue slices. The classic gel recipe results in an expansion factor of ~4×, with a resolution of 60-80 nm. We have recently developed X10 microscopy, which uses a gel that achieves an expansion factor of ~10×, with a resolution of ~25 nm. Here, we provide a step-by-step protocol for X10 expansion microscopy. A typical experiment consists of seven sequential stages: (i) immunostaining, (ii) anchoring, (iii) polymerization, (iv) homogenization, (v) expansion, (vi) imaging, and (vii) validation. The protocol presented here includes recommendations for optimization, pitfalls and their solutions, and detailed guidelines that should increase reproducibility. Although our protocol focuses on X10 expansion microscopy, we detail which of these suggestions are also applicable to classic fourfold expansion microscopy. We exemplify our protocol using primary hippocampal neurons from rats, but our approach can be used with other primary cells or cultured cell lines of interest. This protocol will enable any researcher with basic experience in immunostainings and access to an epifluorescence microscope to perform super-resolution microscopy with X10. The procedure takes 3 d and requires ~5 h of actively handling the sample for labeling and expansion, and another ~3 h for imaging and analysis.
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Registered trials
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