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Novel scanning electron microscopy methods for analyzing the 3D structure of the Golgi apparatus

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Abstract

The structure of the Golgi apparatus has been extensively examined by light and electron microscopy, but details of its three-dimensional (3D) structure have remained unclear because of the technical limitations of conventional microscopy techniques. To overcome this problem, we have developed several novel scanning electron microscopy (SEM) methods for observing the 3D structure of subcellular organelles including the Golgi apparatus: (1) an osmium maceration method that facilitates SEM observation of membranous organelles, including the Golgi apparatus, by selectively removing soluble cytoplasmic proteins, (2) an osmium impregnation/maceration method that combines an osmium impregnation method with the osmium maceration method to determine the polarity of the Golgi apparatus by SEM, (3) a correlative light and SEM method that combines a cryosectioning technique with the osmium maceration method to enable correlation of the immunocytochemical distribution of molecules with the 3D ultrastructure of the Golgi apparatus, and (4) array tomography based on the systematic collection and integration of SEM images of serial ultrathin sections on glass slides for revealing the 3D ultrastructure of the entire Golgi apparatus. Together, the novel SEM techniques listed above can reveal the complete 3D structure of the Golgi apparatus in different cell types.

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Acknowledgements

The authors thank Prof. Emeritus Shohei Yamashina (Department of Anatomy, Kitasato University School of Medicine) and Prof. Emeritus Keiichi Tanaka (Department of Anatomy, Faculty of Medicine, Tottori University) for their mentorship and valuable comments. The authors are also grateful to Drs. Satoshi Kusumi (Division of Morphological Sciences, Kagoshima University Graduate School of Medical and Dental Sciences) and Hiroki Bochimoto (Department of Microscopic Anatomy and Cell Biology, Asahikawa Medical University) for their constructive suggestions and technical support. This work was supported in part by Grants-in-Aid for Young Scientists (B) from the Japanese Society for the Promotion of Science (JSPS) (#21790176 and #26860128), a Grant-in-Aid for Scientific Research (c) from the JSPS (# 16k08458) and a Tsukada Grant for Niigata University Medical Research. This review is based on a lecture given on receipt of the Encouragement Award of the Japanese Association of Anatomists at the 121st annual meeting held at Big Palette, Fukushima, on 29 March 2016.

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  1. Department of Microscopic Anatomy and Cell Biology, Asahikawa Medical University, Midorigaoka Higashi 2-1-1-1, Asahikawa, 078-8510, Japan

    Daisuke Koga & Tsuyoshi Watanabe

  2. Division of Microscopic Anatomy and Bio-imaging, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan

    Daisuke Koga & Tatsuo Ushiki

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  1. Daisuke Koga
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Koga, D., Ushiki, T. & Watanabe, T. Novel scanning electron microscopy methods for analyzing the 3D structure of the Golgi apparatus. Anat Sci Int 92, 37–49 (2017). https://doi.org/10.1007/s12565-016-0380-8

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