A modified ‘NanoSuit®’ preserves wet samples in high vacuum: direct observations on cells and tissues in field-emission scanning electron microscopy

• Main Authors: Yasuharu Takaku, Hiroshi Suzuki, Hideya Kawasaki, Isao Ohta, Daisuke Ishii, Satoshi Hirakawa, Takami Tsutsui, Haruko Matsumoto, Sayuri Takehara, Chinatsu Nakane, Kana Sakaida, Chiaki Suzuki, Yoshinori Muranaka, Hirotoshi Kikuchi, Hiroyuki Konno, Masatsugu Shimomura, Takahiko Hariyama
• Format: Article
• Language: English
• Published: The Royal Society 2017-01-01
• Series: Royal Society Open Science
• Subjects: field-emission scanning electron microscope (fe-sem); high vacuo; living specimen; surface shield effect; nanosuit®
• Online Access: https://royalsocietypublishing.org/doi/pdf/10.1098/rsos.160887

Although field-emission scanning electron microscopy (FE-SEM) has proven very useful in biomedical research, the high vacuum required (10−3 to 10−7 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.