Kubota Yoshiyuki, Sohn Jaerin, Hatada Sayuri, Schurr Meike, Straehle Jakob, Gour Anjali, Neujahr Ralph, Miki Takafumi, Mikula Shawn, Kawaguchi Yasuo
Division of Cerebral Circuitry, National Institute for Physiological Sciences, 5-1 Myodaiji-Higashiyama, Okazaki, Aichi, 444-8787, Japan.
Department of Physiological Sciences, The Graduate University for Advanced Studies (SOKENDAI), 5-1 Myodaiji-Higashiyama, Okazaki, Aichi, 444-8787, Japan.
Nat Commun. 2018 Jan 30;9(1):437. doi: 10.1038/s41467-017-02768-7.
Automated tape-collecting ultramicrotomy in conjunction with scanning electron microscopy (SEM) is a powerful approach for volume electron microscopy and three-dimensional neuronal circuit analysis. Current tapes are limited by section wrinkle formation, surface scratches and sample charging during imaging. Here we show that a plasma-hydrophilized carbon nanotube (CNT)-coated polyethylene terephthalate (PET) tape effectively resolves these issues and produces SEM images of comparable quality to those from transmission electron microscopy. CNT tape can withstand multiple rounds of imaging, offer low surface resistance across the entire tape length and generate no wrinkles during the collection of ultrathin sections. When combined with an enhanced en bloc staining protocol, CNT tape-processed brain sections reveal detailed synaptic ultrastructure. In addition, CNT tape is compatible with post-embedding immunostaining for light and electron microscopy. We conclude that CNT tape can enable high-resolution volume electron microscopy for brain ultrastructure analysis.
结合扫描电子显微镜（SEM）的自动胶带收集超薄切片技术是一种用于体积电子显微镜和三维神经元回路分析的强大方法。目前的胶带存在切片褶皱形成、表面划痕以及成像过程中样品带电等问题。在此，我们表明，经等离子体亲水化处理的碳纳米管（CNT）涂层聚对苯二甲酸乙二醇酯（PET）胶带能有效解决这些问题，并产生与透射电子显微镜质量相当的SEM图像。CNT胶带可承受多轮成像，在整个胶带长度上具有低表面电阻，并且在超薄切片收集过程中不会产生褶皱。当与增强的整体染色方案相结合时，经CNT胶带处理的脑切片可揭示详细的突触超微结构。此外，CNT胶带与用于光学和电子显微镜的包埋后免疫染色兼容。我们得出结论，CNT胶带可实现用于脑超微结构分析的高分辨率体积电子显微镜技术。
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