Geometry-preserving expansion microscopy microplates enable high-fidelity nanoscale distortion mapping

• Published in: Cell reports physical science 2023-12, Vol.4 (12), p.101719, Article 101719
• Main Authors: Seehra, Rajpinder S., Warrington, Samantha J., Allouis, Benjamin H.K., Sheard, Thomas M.D., Spencer, Michael E., Shakespeare, Tayla, Cadby, Ashley, Bose, Daniel, Strutt, David, Jayasinghe, Izzy
• Format: Article
• Language: English
• Subjects: 3D printing; correlative microscopy; distortion mapping; Drosophila melanogaster; expansion microscopy; HeLa cells; multiplexed fluorescence imaging; super-resolution
• ISSN: 2666-3864; 2666-3864
• DOI: 10.1016/j.xcrp.2023.101719

Expansion microscopy (ExM) is a versatile super-resolution microscopy pipeline, leveraging nanoscale biomolecular crosslinking and osmotically driven swelling of hydrogels. Currently, ExM is a laborious and skill-intensive technique, involving manual handling of the hydrogels that can compromise the integrity of the gels and capacity to track gel isotropy, hence diminishing reproducibility. We have developed a 3D-printable microplate system to contain the entire ExM workflow within each well, enabling in situ image acquisition and eliminating the need for direct handling of the hydrogels. The preservation of the gel geometry and orientation of the microplate wells enables convenient tracking of gel expansion, pre- and post-ExM image acquisition, and distortion mapping of every cell or region of interest. We demonstrate the utility of this approach with both single-color and multiplexed ExM of cultured HeLa cells and dissected pupal Drosophila melanogaster wing tissue to reveal distortion-prone structures ranging from sub-cellular organelles to micron-scale tissue regions. [Display omitted] •3D-printed microplate system designed for in situ ExM•Plate-based ExM avoided handling and preserved gel geometry during expansion•Pre- and post-ExM imaging within each well allowed visualization of distortions•Distortions observed in sub-cellular areas in HeLa cells and across fly wing tissue Expansion microscopy is an analytical protocol that allows the physical magnification of biological structures (cells, tissues, and organisms) with swellable hydrogels. Seehra et al. report the development of an array-based approach to containing gel expansion and visualization of spatial distortions intrinsic to magnified cell and tissue ultrastructure.