Dense, Continuous Membrane Labeling and Expansion Microscopy Visualization of Ultrastructure in Tissues.
Dense, Continuous Membrane Labeling and Expansion Microscopy Visualization of Ultrastructure in Tissues.
复制标题
组织中超微结构的密集、连续膜标记和膨胀显微镜可视化。
DOI：
10.1101/2024.03.07.583776
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发表时间：
2024
期刊：
影响因子：
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通讯作者：
IMAXTGrandChallengeConsortium
中科院分区：
文献类型：
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作者：
Shin,TayWon;Wang,Hao;Zhang,Chi;An,Bobae;Lu,Yangning;Zhang,Elizabeth;Lu,Xiaotang;Karagiannis,EmmanouilD;Kang,JeongSeuk;Emenari,Amauche;Symvoulidis,Panagiotis;Asano,Shoh;Lin,Leanne;Costa,EmmaK;IMAXTGrandChallengeConsortium
脂质膜是生物系统的纳米级区室化的关键，但在完整组织中以纳米级精度对其进行荧光可视化，在高标记密度下具有挑战性。在这里，我们报告了超微结构膜扩张显微镜（umExM），它结合了创新的膜标签和优化的扩张显微镜协议，以支持组织中的膜的密集标记，用于纳米级可视化。我们验证了脑切片中umExM膜标记的高信号背景比以及均匀性和连续性，这支持在共聚焦显微镜上以约60 nm的分辨率对膜和蛋白质进行成像。我们证明了umExM的实用程序的神经元过程，如轴突，在小鼠脑组织中的分割和跟踪。将umExM与光学波动成像相结合，或迭代扩展过程，产生约35 nm分辨率的成像，指向在普通光学显微镜上对脑膜进行电子显微镜分辨率可视化的潜力。
Lipid membranes are key to the nanoscale compartmentalization of biological systems, but fluorescent visualization of them in intact tissues, with nanoscale precision, is challenging to do with high labeling density. Here, we report ultrastructural membrane expansion microscopy (umExM), which combines an innovative membrane label and optimized expansion microscopy protocol, to support dense labeling of membranes in tissues for nanoscale visualization. We validate the high signal-to-background ratio, and uniformity and continuity, of umExM membrane labeling in brain slices, which supports the imaging of membranes and proteins at a resolution of ~60 nm on a confocal microscope. We demonstrate the utility of umExM for the segmentation and tracing of neuronal processes, such as axons, in mouse brain tissue. Combining umExM with optical fluctuation imaging, or iterating the expansion process, yields ~35 nm resolution imaging, pointing towards the potential for electron microscopy resolution visualization of brain membranes on ordinary light microscopes.