Brazilian Synchrotron Light National Laboratory, Brazilian Center for Research in Energy and Materials (CNPEM), Giuseppe Maximo Scolfaro 10000, Campinas, São Paulo 13083-970, Brazil.
Brazilian Biosciences National Laboratory, Brazilian Center for Research in Energy and Materials (CNPEM), Giuseppe Maximo Scolfaro 10000, Campinas, São Paulo 13083-970, Brazil.
J Synchrotron Radiat. 2019 Nov 1;26(Pt 6):2013-2023. doi: 10.1107/S1600577519011299. Epub 2019 Sep 10.
The spread of microtomography as a tool for visualization of soft tissues has had a significant impact on a better understanding of complex biological systems. This technique allows a detailed three-dimensional quantitative view of the specimen to be obtained, correlating its morphological organization with its function, providing valuable insights on the functionality of the tissue. Regularly overlooked, but of great importance, proper sample mounting and preparation are fundamental for achieving the highest possible image quality even for the high-resolution imaging systems currently under development. Here, a quantitative analysis compares some of the most common sample-mounting strategies used for synchrotron-based X-ray microtomography of soft tissues: alcoholic-immersion, paraffin-embedding and critical-point drying. These three distinct sample-mounting strategies were performed on the same specimen in order to investigate their impact on sample morphology regardless of individual sample variation. In that sense, the alcoholic-immersion strategy, although causing less shrinkage to the tissue, proved to be the most unsuitable approach for a high-throughput high-resolution imaging experiment due to sample drifting. Also, critical-point drying may present some interesting advantages regarding image quality but is also incompatible with a high-throughput experiment. Lastly, paraffin-embedding is shown to be the most suitable strategy for current soft tissue microtomography experiments. Such detailed analysis of biological sample-mounting strategies for synchrotron-based X-ray microtomography are expected to offer valuable insights on the best approach for using this technique for 3D imaging of soft tissues and following morphometric analysis.
微断层摄影术作为一种软组织可视化工具的传播，对更好地理解复杂的生物系统产生了重大影响。该技术允许获得标本的详细三维定量视图，将其形态组织与其功能相关联，提供有关组织功能的有价值的见解。经常被忽视但非常重要的是，适当的样品安装和准备对于实现即使是目前正在开发的高分辨率成像系统也尽可能高的图像质量是至关重要的。在这里，一种定量分析比较了一些最常见的用于基于同步加速器的软组织 X 射线断层摄影术的样品安装策略：酒精浸泡、石蜡包埋和临界点干燥。这三种不同的样品安装策略在同一标本上进行，以研究它们对样品形态的影响，而不考虑样品的个体变化。从这个意义上说，尽管酒精浸泡策略对组织的收缩较小，但由于样品漂移，它被证明是最不适合高通量高分辨率成像实验的方法。此外，临界点干燥可能在图像质量方面具有一些有趣的优势，但也与高通量实验不兼容。最后，石蜡包埋被证明是当前软组织断层摄影术实验最适合的策略。这种对基于同步加速器的 X 射线断层摄影术的生物样品安装策略的详细分析，有望为该技术在软组织的 3D 成像和形态计量分析方面的最佳应用提供有价值的见解。
