Expansion microscopy of nuclear structure and dynamics in neutrophils

Neutrophils are key players of the immune system and possess an arsenal of effector functions, including the ability to form and expel neutrophil extracellular traps (NETs) in a process termed NETosis. During NETosis, the nuclear DNA/chromatin expands until it fills the whole cell and is released into the extracellular space. NETs are composed of DNA decorated with histones, proteins or peptides and NETosis is implicated in many diseases. Resolving the structure and dynamics of the nucleus in great detail is essential to understand the underlying processes but so far super-resolution methods have not been applied. Here, we developed an expansion microscopy-based method and determined the spatial distribution of chromatin/DNA, histone H1, and nucleophosmin (NPM1) with a 4.9-fold improved resolution (< 40 nm) and increased information content. It allowed us to identify the punctate localization of NPM1 in the nucleus and histone-rich domains in NETotic cells with a size of 54 nm. The technique could also be applied to components of the nuclear envelope (lamins B1 and B2) and myeloperoxidase (MPO) providing a complete picture of nuclear dynamics and structure. In conclusion, expansion microscopy enables super-resolved imaging of the highly dynamic structure of nuclei in immune cells. Why it matters Accessibility to high-resolution imaging is critical to advancing research across various disciplines. However, conventionally this requires demanding optical hardware, special fluorophores or data analysis. Expansion microscopy is a technique adaptable to different cell and tissue types and is comparatively inexpensive and easy to perform. Applying this technique to cells and compartments such as the nucleus of immune cells that are difficult to image due to their size and morphology, yields valuable structural insights that would otherwise require more difficult super-resolution methods. ### Competing Interest Statement The authors have declared no competing interest.