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

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• 作者：Daisuke Koga ; Tatsuo Ushiki ; Tsuyoshi Watanabe
• 关键词：3D ; Array tomography using SEM ; Golgi apparatus ; Osmium maceration method ; SEM
• 刊名：Anatomical Science International
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：92
• 期：1
• 页码：37-49
• 全文大小：
• 刊物主题：Anatomy; Human Physiology; Neurosciences; Cell Biology; Animal Anatomy / Morphology / Histology; Animal Physiology;
• 出版者：Springer Japan
• ISSN：1447-073X
• 卷排序：92

文摘

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.