Takaku Yasuharu, Suzuki Hiroshi, Kawasaki Hideya, Ohta Isao, Ishii Daisuke, Hirakawa Satoshi, Tsutsui Takami, Matsumoto Haruko, Takehara Sayuri, Nakane Chinatsu, Sakaida Kana, Suzuki Chiaki, Muranaka Yoshinori, Kikuchi Hirotoshi, Konno Hiroyuki, Shimomura Masatsugu, Hariyama Takahiko
Department of Biology , Hamamatsu University School of Medicine , Higashi-ku, Hamamatsu 431-3192 , Japan.
Department of Chemistry , Hamamatsu University School of Medicine , Higashi-ku, Hamamatsu 431-3192 , Japan.
R Soc Open Sci. 2017 Mar 1;4(3):160887. doi: 10.1098/rsos.160887. eCollection 2017 Mar.
Although field-emission scanning electron microscopy (FE-SEM) has proven very useful in biomedical research, the high vacuum required (10 to 10 Pa) precludes direct observations of living cells and tissues at high resolution and often produces unwanted structural changes. We have previously described a method that allows the investigator to keep a variety of insect larvae alive in the high vacuum environment of the electron microscope by encasing the organisms in a thin, vacuum-proof suit, the 'NanoSuit®'. However, it was impossible to protect wet tissues freshly excised from intact organisms or cultured cells. Here we describe an improved 'NanoSuit' technique to overcome this limitation. We protected the specimens with a surface shield enhancer (SSE) solution that consists of glycerine and electrolytes and found that the fine structure of the SSE-treated specimens is superior to that of conventionally prepared specimens. The SSE-based NanoSuit affords a much stronger barrier to gas and/or liquid loss than the previous NanoSuit did and, since it allows more detailed images, it could significantly help to elucidate the 'real' organization of cells and their functions.
尽管场发射扫描电子显微镜（FE-SEM）在生物医学研究中已被证明非常有用，但所需的高真空环境（10至10 Pa）排除了对活细胞和组织进行高分辨率的直接观察，并且常常会产生不必要的结构变化。我们之前描述了一种方法，通过将生物体包裹在一种薄的、防真空的“纳米防护服（NanoSuit®）”中，使研究者能够在电子显微镜的高真空环境中让各种昆虫幼虫存活。然而，对于从完整生物体或培养细胞中新鲜切除的湿组织，却无法进行保护。在此，我们描述一种改进的“纳米防护服”技术以克服这一限制。我们用一种由甘油和电解质组成的表面屏蔽增强剂（SSE）溶液来保护标本，发现经SSE处理的标本的精细结构优于传统制备的标本。基于SSE的纳米防护服比之前的纳米防护服对气体和/或液体损失提供了更强的屏障，并且由于它能提供更详细的图像，故能显著有助于阐明细胞的“真实”组织结构及其功能。
