Micheva Kristina D, O'Rourke Nancy, Busse Brad, Smith Stephen J
Cold Spring Harb Protoc. 2010 Nov 1;2010(11):pdb.prot5523. doi: 10.1101/pdb.prot5523.
Array tomography is a volumetric microscopy method based on physical serial sectioning. Ultrathin sections of a plastic-embedded tissue are cut using an ultramicrotome, bonded in an ordered array to a glass coverslip, stained as desired, and imaged. The resulting two-dimensional image tiles can then be reconstructed computationally into three-dimensional volume images for visualization and quantitative analysis. The minimal thickness of individual sections permits high-quality rapid staining and imaging, whereas the array format allows reliable and convenient section handling, staining, and automated imaging. Also, the physical stability of the arrays permits images to be acquired and registered from repeated cycles of staining, imaging, and stain elution, as well as from imaging using multiple modalities (e.g., fluorescence and electron microscopy). Array tomography makes it possible to visualize and quantify previously inaccessible features of tissue structure and molecular architecture. However, careful preparation of the tissue is essential for successful array tomography; these steps can be time-consuming and require some practice to perfect. This protocol describes the fixation and processing required to prepare tissues for immunofluorescence array tomography.
阵列断层扫描是一种基于物理连续切片的体视显微镜方法。使用超薄切片机切割塑料包埋组织的超薄切片，将其按有序阵列粘贴到玻璃盖玻片上，根据需要进行染色并成像。然后可以通过计算将所得的二维图像切片重建为三维体积图像，以进行可视化和定量分析。单个切片的最小厚度允许进行高质量的快速染色和成像，而阵列形式则允许可靠且方便地处理切片、染色和自动成像。此外，阵列的物理稳定性允许从染色、成像和洗脱染色的重复循环以及使用多种模式（例如荧光和电子显微镜）成像中获取和配准图像。阵列断层扫描使得可视化和量化以前无法访问的组织结构和分子结构特征成为可能。然而，仔细准备组织对于成功进行阵列断层扫描至关重要；这些步骤可能很耗时，并且需要一些实践才能完善。本方案描述了为免疫荧光阵列断层扫描准备组织所需的固定和处理方法。
