title: Observation of tissues in open aqueous solution by atmospheric scanning electron microscopy: applicability to intraoperative cancer diagnosis.
abstract: In the atmospheric scanning electron microscope (ASEM), a 2- to 3-µm layer of the sample resting on a silicon nitride-film window in the base of an open sample dish is imaged, in liquid, at atmospheric pressure, from below by an inverted SEM. Thus, the time-consuming pretreatments generally required for biological samples to withstand the vacuum of a standard electron microscope are avoided. In the present study, various mouse tissues (brain, spinal cord, muscle, heart, lung, liver, kidney, spleen and stomach) were fixed, stained with heavy metals, and visualized in radical scavenger D-glucose solution using the ASEM. While some stains made the nuclei of cells very prominent (platinum-blue, phosphotungstic acid), others also emphasized cell organelles and membranous structures (uranium acetate or the NCMIR method). Notably, symbiotic bacteria were sometimes observed on stomach mucosa. Furthermore, kidney tissue could be stained and successfully imaged in <30 min. Lung and spinal cord tissue from normal mice and mice metastasized with breast cancer cells was also examined. Cancer cells present in lung alveoli and in parts of the spine tissue clearly had larger nuclei than normal cells. The results indicate that the ASEM has the potential to accelerate intraoperative cancer diagnosis, the diagnosis of kidney diseases and pathogen detection. Importantly, in the course of the present study it was possible to increase the observable tissue area by using a new multi-windowed ASEM sample dish and sliding the tissue across its eight windows.
authors.affiliation: Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-0006, Japan.
authors.forename: Nassirhadjy
authors.initials: N
authors.lastname: Memtily
authors.affiliation: Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8568, Japan.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Okada
authors.affiliation: Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8568, Japan.
authors.forename: Tatsuhiko
authors.initials: T
authors.lastname: Ebihara
authors.affiliation: Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8568, Japan.
authors.forename: Mari
authors.initials: M
authors.lastname: Sato
authors.affiliation: Department of Pathology, Kochi Medical School, University of Kochi, Nankoku, Kochi 783-8505, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Kurabayashi
authors.affiliation: Department of Pathology, Kochi Medical School, University of Kochi, Nankoku, Kochi 783-8505, Japan.
authors.forename: Mutsuo
authors.initials: M
authors.lastname: Furihata
authors.affiliation: Advanced Technology Division, JEOL Ltd., Akishima, Tokyo 196‑8558, Japan.
authors.forename: Mitsuo
authors.initials: M
authors.lastname: Suga
authors.affiliation: Advanced Technology Division, JEOL Ltd., Akishima, Tokyo 196‑8558, Japan.
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Nishiyama
authors.affiliation: Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-0006, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Mio
authors.affiliation: Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-0006, Japan.
authors.forename: Chikara
authors.initials: C
authors.lastname: Sato
mesh_terms: D000818:Animals; D001274:Atmospheric Pressure; D005260:Female; D007430:Intraoperative Care; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008813:Mice, Inbred ICR; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D009369:Neoplasms / Q000175:diagnosis* / Q000601:surgery; D013194:Staining and Labeling / Q000379:methods