Three-Dimensional Microscopy by Milling with Ultraviolet Excitation
Three-Dimensional Microscopy by Milling with Ultraviolet Excitation
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DOI：
10.1038/s41598-019-50870-1
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发表时间：
2019-10-10
影响因子：
4.6
通讯作者：
Mayerich, David
中科院分区：
文献类型：
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作者：
Guo, Jiaming;Artur, Camille;Mayerich, David
三维生物样本的分析对于了解组织功能和疾病机制至关重要。许多慢性疾病，如神经退行性疾病和癌症，与复杂的组织变化相关，难以使用二维组织学进行探索。虽然三维技术，如共聚焦和光片显微镜是完善的，他们是耗时的，需要昂贵的仪器，并限于小组织体积。因此，三维显微镜在临床环境中是不切实际的，并且通常仅限于主要研究机构的核心设施。为临床医生和研究人员提供以细胞分辨率对大型三维组织体积进行常规成像的能力将有巨大的好处。在本文中，我们提出了一种成像方法，使快速和廉价的三维成像，可以很容易地集成到当前的组织学管道。该方法依赖于使用深紫外激发的石蜡包埋样品的块面成像。然后烧蚀成像表面以显露下一个用于成像的组织切片。最终的图像堆栈然后被对齐和重建，以提供超过现代三维成像系统可实现的深度和分辨率的组织模型。
Analysis of three-dimensional biological samples is critical to understanding tissue function and the mechanisms of disease. Many chronic conditions, like neurodegenerative diseases and cancers, correlate with complex tissue changes that are difficult to explore using two-dimensional histology. While three-dimensional techniques such as confocal and light-sheet microscopy are well-established, they are time consuming, require expensive instrumentation, and are limited to small tissue volumes. Three-dimensional microscopy is therefore impractical in clinical settings and often limited to core facilities at major research institutions. There would be a tremendous benefit to providing clinicians and researchers with the ability to routinely image large three-dimensional tissue volumes at cellular resolution. In this paper, we propose an imaging methodology that enables fast and inexpensive three-dimensional imaging that can be readily integrated into current histology pipelines. This method relies on block-face imaging of paraffin-embedded samples using deep-ultraviolet excitation. The imaged surface is then ablated to reveal the next tissue section for imaging. The final image stack is then aligned and reconstructed to provide tissue models that exceed the depth and resolution achievable with modern three-dimensional imaging systems.