Protocols
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Explore our Expansion Microscopy developments and applications
Beyond the protocols presented here, our lab has developed and optimized several Expansion Microscopy approaches to investigate cellular architecture at the nanoscale. Discover our methodological developments, practical applications, and examples from our research on our dedicated page:
👉 Explore our Expansion Microscopy developments and applications
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1- Getting Started with Cryo-ExM & U-ExM
Cryo-ExM equipment: Manual Plunge Freezers
Cryo-ExM requires rapid sample vitrification, which can be achieved using automated plunge freezers, such as the Vitrobot, or simpler, more affordable manual plunge-freezing systems.
In our laboratory, we use manual plunge-freezing systems from the following manufacturers:
Both manufacturers offer equipment suitable for plunge-freezing workflows. We recommend contacting them directly to discuss the most appropriate configuration for your samples.
2. U-ExM across biological systems: protocols and resources from the community
Ultrastructure Expansion Microscopy (U-ExM) has been successfully adapted to a wide range of biological systems, from cultured cells to complex tissues and diverse organisms. Here, we highlight a growing collection of protocols and methodological adaptations developed by our friends and colleagues across the microscopy community.
Whether you're working with mammalian tissues, model organisms, or more specialized biological systems, these resources may help you adapt U-ExM to your own research.
2a. Kidney tissues — Mouse and human
Langner et al., Cytoskeleton (2024).
A detailed, step-by-step adaptation of U-ExM for mouse and human kidney sections, enabling visualization of centrioles, cilia, cytoskeletal structures, and basement membranes. Compatible with different fixation and embedding conditions.
2b. Drosophila — Cultured cells, testes, larval brains and embryos
1. Visualizing Drosophila centrioles by expansion microscopy
A detailed, step-by-step expansion microscopy protocol adapted for Drosophila S2 cultured cells and tissues, including testes and ovaries. The study demonstrates applications to centriole architecture, procentriole formation, and protein localization, with practical guidance for sample preparation, expansion, and imaging.
Brunet et al., The EMBO Journal (2025).
An adaptation of U-ExM to several Drosophila tissues, including testes, larval brains, and early embryos. The approach enables nanoscale mapping of centriolar proteins and was used to reveal the distinct localization patterns of Alms1a and Alms1b during centriole duplication.
2c. Apicomplexan parasites — Plasmodium, Toxoplasma and Cryptosporidium
Expansion microscopy of apicomplexan parasites
Liffner & Absalon, Molecular Microbiology (2024) — Review article.
An excellent overview of U-ExM applications in apicomplexan parasites. The review covers Plasmodium, Toxoplasma, and Cryptosporidium, with examples of its use to investigate cell division, cytoskeletal organization, organelle architecture, and protein localization across different life-cycle stages.
It also discusses emerging ExM adaptations and future applications in parasitology.
2d. Mosquito tissues — Malaria parasites and host–parasite interactions
A specialized adaptation of U-ExM for mosquito tissues, enabling high-resolution visualization of Plasmodium parasites within their mosquito host.
MoTissU-ExM facilitates the investigation of parasite ultrastructure, tissue architecture, and host–parasite interactions during malaria transmission.
2e. Toxoplasma gondii — Enzymatically enhanced U-ExM for tissue cysts
Bondarenko et al., mSphere (2024).
An adaptation of U-ExM specifically developed for the chronic cyst-forming stage of Toxoplasma gondii.
By introducing an enzymatic digestion step using the mucinase StcE, this protocol overcomes the resistance of the cyst wall to conventional expansion, enabling approximately 4× expansion and nanoscale visualization of parasite structures and protein organization within cysts grown in fibroblast and neuronal cultures.
2f. Canine retina — Photoreceptor architecture
A detailed U-ExM adaptation for fresh and archival canine retinal tissues, including frozen sections.
The article provides extensive methodological information and supplementary protocols for examining photoreceptor connecting cilia, calyceal processes, and protein localization.
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3- Additional Expansion Microscopy Protocols and Resources
Looking for more ExM protocols, practical tips, or troubleshooting advice? In 2021, we edited the book Expansion Microscopy for Cell Biology, which brings together a comprehensive collection of protocols covering different ExM approaches. It provides a valuable resource for researchers looking to explore, optimize, or implement expansion microscopy techniques in their laboratories.
https://shop.elsevier.com/books/expansion-microscopy-for-cell-biology/guichard/978-0-12-820807-6
