Corrected Super-Resolution Microscopy Enables Nanoscale Imaging of Autofluorescent Lung Macrophages.
Corrected Super-Resolution Microscopy Enables Nanoscale Imaging of Autofluorescent Lung Macrophages.
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DOI：
10.1016/j.bpj.2020.10.041
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发表时间：
2020-12-15
影响因子：
3.4
通讯作者：
Davis DM
中科院分区：
文献类型：
--
作者：
Ambrose AR;Dechantsreiter S;Shah R;Montero MA;Quinn AM;Hessel EM;Beinke S;Tannahill GM;Davis DM
使用超分辨率显微镜以纳米级分辨率观察细胞表面和底层细胞骨架，使人们能够深入了解细胞信号传导和功能。然而，组织特异性免疫细胞的纳米级动力学研究相对较少。例如，组织巨噬细胞具有高度自发荧光，严重限制了光学显微镜的实用性。在这里，我们报告了一种从随机光学重建显微镜（STORM）数据集中消除自发荧光噪声的校正技术。实验数据的模拟和分析表明，移动中值滤波器是一种准确而稳健的校正技术，可广泛应用于具有挑战性的生物样本。在这里，我们使用这种方法通过抗体包被的载玻片来可视化通过 Fc 受体激活的肺巨噬细胞。对巨噬细胞形态的精确、纳米级定量表明，激活诱导了末端带有 MHC I 类蛋白的细胞突起的形成。这些数据与肺巨噬细胞突起在抗原呈递中的作用一致。此外，已知标记细胞外囊泡的四跨膜蛋白 CD81 在活化的肺巨噬细胞表面以环形结构（平均直径 93 ± 50 nm）出现。因此，移动中值滤波器校正技术使我们能够定量分析组织源性免疫细胞的细胞外分泌物和膜结构。
Observing the cell surface and underlying cytoskeleton at nanoscale resolution using super-resolution microscopy has enabled many insights into cell signaling and function. However, the nanoscale dynamics of tissue-specific immune cells have been relatively little studied. Tissue macrophages, for example, are highly autofluorescent, severely limiting the utility of light microscopy. Here, we report a correction technique to remove autofluorescent noise from stochastic optical reconstruction microscopy (STORM) data sets. Simulations and analysis of experimental data identified a moving median filter as an accurate and robust correction technique, which is widely applicable across challenging biological samples. Here, we used this method to visualize lung macrophages activated through Fc receptors by antibody-coated glass slides. Accurate, nanoscale quantification of macrophage morphology revealed that activation induced the formation of cellular protrusions tipped with MHC class I protein. These data are consistent with a role for lung macrophage protrusions in antigen presentation. Moreover, the tetraspanin protein CD81, known to mark extracellular vesicles, appeared in ring-shaped structures (mean diameter 93 ± 50 nm) at the surface of activated lung macrophages. Thus, a moving median filter correction technique allowed us to quantitatively analyze extracellular secretions and membrane structure in tissue-derived immune cells.
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影响因子：
7.3
作者：
Andreu Z;Yáñez-Mó M
通讯作者：
Yáñez-Mó M
DOI：
10.1126/science.aau6977
发表时间：
2020-02-07
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Kalluri R;LeBleu VS
通讯作者：
LeBleu VS
影响因子：
8.4
作者：
Jun YW;Kim HR;Reo YJ;Dai M;Ahn KH
通讯作者：
Ahn KH
影响因子：
7.3
作者：
Bálint Š;Lopes FB;Davis DM
通讯作者：
Davis DM
影响因子：
5.8
作者：
Hsiao, Chia-Hung Christine;Ueno, Norikiyo;Wilson, Mary E.
通讯作者：
Wilson, Mary E.