Robarts Research Institute, The University of Western Ontario, London, Ontario, N6A 5B7, Canada.
Department of Medical Biophysics, The University of Western Ontario, London, Ontario, N6A 5C1, Canada.
Sci Rep. 2019 Jan 24;9(1):698. doi: 10.1038/s41598-018-36905-z.
Virtual histology - utilizing high-resolution three-dimensional imaging - is becoming readily available. Micro-computed tomography (micro-CT) is widely available and is often coupled with x-ray attenuating histological stains that mark specific tissue components for 3D virtual histology. In this study we describe a new tri-element x-ray attenuating stain and perfusion protocol that provides micro-CT contrast of the entire vasculature of an intact mouse. The stain - derived from an established histology stain (Verhoeff's) - is modified to enable perfusion through the vasculature; the attenuating elements of the stain are iodine, aluminum, and iron. After a 30-minute perfusion through the vasculature (10-minute flushing with detergent-containing saline followed by 15-minute perfusion with the stain and a final 5-minute saline flush), animals are scanned using micro-CT. We demonstrate that the new staining protocol enables sharp delineation of the vessel walls in three dimensions over the whole body; corresponding histological analysis verified that the CT stain is localized primarily in the endothelial cells and media of large arteries and the endothelium of smaller vessels, such as the coronaries. The rapid perfusion and scanning protocol ensured that all tissues are available for further analysis via higher resolution CT of smaller sections or traditional histological sectioning.
虚拟组织学——利用高分辨率三维成像——正变得易于获得。微计算机断层扫描（micro-CT）广泛可用，并且通常与 X 射线衰减组织学染色结合使用，这些染色标记特定的组织成分以进行 3D 虚拟组织学。在这项研究中，我们描述了一种新的三元素 X 射线衰减染色和灌注方案，该方案可提供完整小鼠整个脉管系统的微 CT 对比。该染色剂——源自一种已建立的组织学染色剂（Verhoeff's）——经过改良后可使血管内的血液灌注；染色剂的衰减元素是碘、铝和铁。在血管内进行 30 分钟的灌注（用含洗涤剂的盐水冲洗 10 分钟，然后用染色剂灌注 15 分钟，最后用盐水冲洗 5 分钟）后，使用 micro-CT 对动物进行扫描。我们证明，新的染色方案能够在整个身体的三维空间中清晰地描绘血管壁；相应的组织学分析证实，CT 染色主要定位于大动脉的内皮细胞和中膜以及较小血管（如冠状动脉）的内皮细胞中。快速的灌注和扫描方案确保了所有组织都可通过较小切片的更高分辨率 CT 或传统的组织学切片进行进一步分析。
