Department of Molecular Cell Biology, Leiden University Medical Center, 2300 RC Leiden, Netherlands.
J Cell Biol. 2012 Aug 6;198(3):457-69. doi: 10.1083/jcb.201201140.
A key obstacle in uncovering the orchestration between molecular and cellular events is the vastly different length scales on which they occur. We describe here a methodology for ultrastructurally mapping regions of cells and tissue as large as 1 mm(2) at nanometer resolution. Our approach employs standard transmission electron microscopy, rapid automated data collection, and stitching to create large virtual slides. It greatly facilitates correlative light-electron microscopy studies to relate structure and function and provides a genuine representation of ultrastructural events. The method is scalable as illustrated by slides up to 281 gigapixels in size. Here, we applied virtual nanoscopy in a correlative light-electron microscopy study to address the role of the endothelial glycocalyx in protein leakage over the glomerular filtration barrier, in an immunogold labeling study of internalization of oncolytic reovirus in human dendritic cells, in a cryo-electron microscopy study of intact vitrified mouse embryonic cells, and in an ultrastructural mapping of a complete zebrafish embryo slice.
揭示分子和细胞事件之间的协调作用的一个主要障碍是它们发生的长度尺度大不相同。我们在这里描述了一种方法，用于以纳米分辨率对 1 平方毫米（2）大小的细胞和组织区域进行超微结构测绘。我们的方法采用标准的透射电子显微镜、快速自动数据采集和拼接来创建大型虚拟幻灯片。它极大地促进了相关的光电子显微镜研究，以关联结构和功能，并提供超微结构事件的真实表示。该方法是可扩展的，如图所示，幻灯片的大小高达 281 吉像素。在这里，我们在一项相关的光电子显微镜研究中应用了虚拟纳米显微镜来解决内皮糖萼在肾小球滤过屏障上的蛋白质渗漏中的作用，在一项关于溶瘤单纯疱疹病毒在人树突状细胞内化的免疫金标记研究中，在一项完整的冷冻电子显微镜研究中在小鼠胚胎细胞中，以及在完整的斑马鱼胚胎切片的超微结构测绘中。
