BACKGROUND Dense and unbiased cellular-resolution representations of extended volumetric central nervous system soft-tissue anatomy are difficult to obtain, even in experimental post-mortem settings. Interestingly, X-ray phase-contrast computed tomography (X-PCI-CT), an emerging soft-tissue-sensitive volumetric imaging technique, can provide multiscale organ- to cellular-level morphological visualizations of neuroanatomical structure. NEW METHOD Here, we tested different nervous-tissue fixation procedures, conventionally used for transmission electron microscopy, to better establish X-PCI-CT-specific sample-preparation protocols. Extracted rat spinal medullas were alternatively fixed with a standard paraformaldehyde-only aldehyde-based protocol, or in combination with glutaraldehyde. Some specimens were additionally post-fixed with osmium tetroxide. Multiscale X-PCI-CT datasets were collected at several synchrotron radiation facilities, using state-of-the-art setups with effective image voxel sizes of 3.03 to 0.33 μm3, and compared to high-field magnetic resonance imaging, histology and vascular fluorescence microscopy data. RESULTS Multiscale X-PCI-CT of aldehyde-fixed spinal cord specimens resulted in dense histology-like volumetric representations and quantifications of extended deep spinal micro-vascular networks and of intra-medullary cell populations. Osmium post-fixation increased intra-medullary contrast between white and gray-matter tissues, and enhanced delineation of intra-medullary cellular structure, e.g. axon fibers and motor neuron perikarya. COMPARISON WITH EXISTING METHODS Volumetric X-PCI-CT provides complementary contrast and higher spatial resolution compared to 9.4 T MRI. X-PCI-CT's advantage over planar histology is the volumetric nature of the cellular-level data obtained, using samples much larger than those fit for volumetric vascular fluorescence microscopy. CONCLUSIONS Deliberately choosing (post-)fixation protocols tailored for optimal nervous-tissue structural preservation is of paramount importance in achieving effective and targeted neuroimaging via the X-PCI-CT technique.

中文翻译：

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建立啮齿动物脊髓的X射线相衬CT的样品制备方案：醛固定和浸渍。

背景技术即使在实验验尸设置中，也很难获得扩展的中枢神经系统软组织解剖结构的密集且无偏差的细胞分辨率表示。有趣的是，X射线相衬计算机断层扫描（X-PCI-CT）是一种新兴的对软组织敏感的体积成像技术，可以为神经解剖结构提供多尺度的器官到细胞水平的形态学可视化。新方法在这里，我们测试了常规用于透射电子显微镜的不同神经组织固定程序，以更好地建立X-PCI-CT特定的样品制备方案。或者，将提取的大鼠脊髓髓质用标准的仅基于低聚甲醛的醛固定，或与戊二醛联合固定。一些标本还用四氧化进行后固定。在多个同步加速器辐射设施收集了多尺度X-PCI-CT数据集，使用了具有3.03至0.33μm3的有效图像体素大小的最新设置，并与高场磁共振成像，组织学和血管荧光显微镜进行了比较数据。结果醛固定脊髓标本的多尺度X-PCI-CT产生了密集的组织学样体积表示，并对扩展的深部脊髓微血管网络和髓内细胞群进行了定量。s固定后增加了白色和灰色物质组织之间的髓内对比度，并增强了髓内细胞结构（如轴突纤维和运动神经元周围核）的轮廓。与现有方法的比较与9.4 T MRI相比，体积X-PCI-CT可提供互补的对比度和更高的空间分辨率。X-PCI-CT相对于平面组织学的优势是所获得的细胞水平数据的体积性质，所使用的样品要比适合于体积血管荧光显微镜的样品大得多。结论故意选择针对最佳神经组织结构保存量身定制的（后）固定方案，对于通过X-PCI-CT技术实现有效且有针对性的神经成像至关重要。