Department of Biotechnology and Biophysics, Theodor-Boveri-Institute, Biocenter, Julius Maximilian University, Würzburg, Germany.
Department of Internal Medicine II, WÜ4i, University Hospital Würzburg, Würzburg, Germany.
Commun Biol. 2021 Oct 4;4(1):1151. doi: 10.1038/s42003-021-02669-y.
Expansion microscopy (ExM) enables super-resolution fluorescence imaging on standard microscopes by physical expansion of the sample. However, the investigation of interactions between different organisms such as mammalian and fungal cells by ExM remains challenging because different cell types require different expansion protocols to ensure identical, ideally isotropic expansion of both partners. Here, we introduce an ExM method that enables super-resolved visualization of the interaction between NK cells and Aspergillus fumigatus hyphae. 4-fold expansion in combination with confocal fluorescence imaging allows us to resolve details of cytoskeleton rearrangement as well as NK cells' lytic granules triggered by contact with an RFP-expressing A. fumigatus strain. In particular, subdiffraction-resolution images show polarized degranulation upon contact formation and the presence of LAMP1 surrounding perforin at the NK cell-surface post degranulation. Our data demonstrate that optimized ExM protocols enable the investigation of immunological synapse formation between two different species with so far unmatched spatial resolution.
扩展显微镜 (ExM) 通过对样品的物理扩展，使标准显微镜能够实现超分辨率荧光成像。然而，通过 ExM 研究不同生物体（如哺乳动物和真菌细胞）之间的相互作用仍然具有挑战性，因为不同的细胞类型需要不同的扩展方案来确保两个伙伴的相同、理想的各向同性扩展。在这里，我们介绍了一种 ExM 方法，该方法能够对 NK 细胞与烟曲霉菌丝体之间的相互作用进行超分辨可视化。4 倍的扩展结合共聚焦荧光成像，使我们能够解析细胞骨架重排的细节以及与表达 RFP 的烟曲霉菌株接触后 NK 细胞的溶酶体颗粒的触发。特别是，亚衍射分辨率的图像显示在接触形成时极化的脱粒，以及在脱粒后 LAMP1 围绕穿孔素存在于 NK 细胞表面。我们的数据表明，优化的 ExM 方案能够以迄今为止无与伦比的空间分辨率研究两种不同物种之间的免疫突触形成。
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