Quantitative 3D imaging of whole, unstained cells by using X-ray diffraction microscopy
Quantitative 3D imaging of whole, unstained cells by using X-ray diffraction microscopy
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DOI：
10.1073/pnas.1000156107
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发表时间：
2010-06-22
影响因子：
11.1
通讯作者：
Miao, Jianwei
中科院分区：
文献类型：
--
作者：
Jiang, Huaidong;Song, Changyong;Miao, Jianwei
显微镜极大地促进了我们对生物学的理解。虽然最近在光学显微镜方面取得了重大进展，打破了衍射极限障碍，但这种技术对荧光标记技术的依赖禁止了对细胞全部内容物的定量3D成像。冷冻电子显微镜可以以3-5 nm的分辨率对多形性结构成像，但仅适用于薄的或切片的标本。在这里，我们报告了一个完整的，未染色的细胞在50-60 nm的分辨率通过X射线衍射显微镜定量三维成像。我们确定了酵母孢子细胞内细胞器的3D形态和结构，包括细胞壁、液泡、内质网、线粒体、颗粒、细胞核和核仁。此外，我们观察到一个三维结构突出的重组酵母孢子，这表明孢子萌发过程。使用低温技术，通过X射线衍射显微镜应该可以实现5-10 nm的3D分辨率。因此，这项工作为以纳米级分辨率对各种生物标本进行定量3D成像铺平了道路，这些生物标本对于电子显微镜来说太厚了。
Microscopy has greatly advanced our understanding of biology. Although significant progress has recently been made in optical microscopy to break the diffraction-limit barrier, reliance of such techniques on fluorescent labeling technologies prohibits quantitative 3D imaging of the entire contents of cells. Cryoelectron microscopy can image pleomorphic structures at a resolution of 3-5 nm, but is only applicable to thin or sectioned specimens. Here, we report quantitative 3D imaging of a whole, unstained cell at a resolution of 50-60 nm by X-ray diffraction microscopy. We identified the 3D morphology and structure of cellular organelles including cell wall, vacuole, endoplasmic reticulum, mitochondria, granules, nucleus, and nucleolus inside a yeast spore cell. Furthermore, we observed a 3D structure protruding from the reconstructed yeast spore, suggesting the spore germination process. Using cryogenic technologies, a 3D resolution of 5-10 nm should be achievable by X-ray diffraction microscopy. This work hence paves a way for quantitative 3D imaging of a wide range of biological specimens at nanometer-scale resolutions that are too thick for electron microscopy.