Three-dimensional observation of foraminiferal cytoplasmic morphology and internal structures using uranium-osmium staining and micro-X-ray computed tomography
Three-dimensional observation of foraminiferal cytoplasmic morphology and internal structures using uranium-osmium staining and micro-X-ray computed tomography
复制标题
DOI：
10.1016/j.marmicro.2015.09.003
复制
发表时间：
2015-12-01
影响因子：
1.9
通讯作者：
Tame, Akihiro
中科院分区：
文献类型：
--
作者：
Nomaki, Hidetaka;Toyofuku, Takashi;Tame, Akihiro
底栖有孔虫是常见的原生生物，栖息在海底广阔的环境范围内。由于其巨大的生物量，这些生物体在生物地球化学循环中发挥着至关重要的作用，但我们对其生态学和细胞生物学的了解仍然有限。原因之一是有孔虫的钙质或凝集测试（壳）妨碍从生物体外部观察或测量细胞质。在这里，我们报告了使用显微 X 射线计算机断层扫描 (CT) 以及基于锇和铀的细胞质染色方法（透射电子显微镜 (TEM) 观察的标准方案）的组合技术对深海底栖有孔虫的细胞质和几种超微结构进行的三维 (3-D) 观察。锇和铀与有机膜、蛋白质和核酸结合，由于其高 X 射线衰减，使得它们可以通过 X 射线可见，并能够将它们重建为 3D 图像。这种方法不仅揭示了外部细胞形态，还揭示了细胞内部的液泡和栓塞。此外，可以使用获得的 CT 数据对细胞质和液泡进行体积计算，以评估其生物量和液泡的作用。我们的方法提供了对整个单个或多个有孔虫细胞的快速（在本研究中不到一个小时）评估，并且可用于研究有孔虫和其他有睾丸真核生物的生物学和生态学，其内部特征被其外部测试结构掩盖。 (C) 2015 Elsevier B.V. 保留所有权利。
Benthic foraminifera are common protists that inhabit a broad environmental range of the seafloor. Because of their large biomass, these organisms play crucial roles in biogeochemical cycles, but our knowledge of their ecology and cell biology are still limited. One reason is that the calcareous or agglutinated tests (shells) of foraminifera hamper observing or measuring the cytoplasm from outside the organism. Here, we report the three-dimensional (3-D) observation of the cytoplasm and several ultrastructures of deep-sea benthic foraminifera using the combined techniques of micro-X-ray computed tomography (CT) and the osmium- and uranium-based cytoplasm-staining methods that are standard protocols for transmission electron microscope (TEM) observation. Osmium and uranium bind to organic membranes, proteins, and nucleic acids, rendering them visible by X-rays due to their high X-ray attenuation and enabling their reconstruction as 3-D images. This methodology revealed not only the external cellular morphology but also vacuoles and plugs inside the cell. Furthermore, volumetric calculation of cytoplasm and vacuoles is possible using the obtained CT data to evaluate their biomass and roles of vacuoles. Our method offers rapid (less than an hour in this study) evaluation of whole single or multiple foraminiferal cells and can be applied to investigate the biology and ecology of the foraminifera and other testate eukaryotes whose internal features are obscured by their external test structures. (C) 2015 Elsevier B.V. All rights reserved.