Advanced CUBIC tissue clearing for whole-organ cell profiling
Advanced CUBIC tissue clearing for whole-organ cell profiling
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DOI：
10.1038/s41596-019-0240-9
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发表时间：
2019-12-01
期刊：
影响因子：
14.8
通讯作者：
Ueda, Hiroki R.
中科院分区：
文献类型：
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作者：
Matsumoto, Katsuhiko;Mitani, Tomoki T.;Ueda, Hiroki R.
组织清除技术是生物学研究和病理诊断的有力工具。在这里，我们描述了先进的清晰、畅通无阻的脑成像鸡尾酒和计算分析(立方体)程序，可以应用于生物医学研究。这种方法可以制备高透明度的器官，通过浸泡在立方体试剂中在7-21天内保留荧光蛋白质信号。然后，透明的小鼠器官可以用高速成像系统(0.5TB/h/色彩)成像。此外，为了更好地理解和简化数据的处理，可以在3-12小时内从大型图像数据集(2.5-14 TB)中提取一个器官(3-12 GB)中所有检测到的细胞的位置。作为如何使用该协议的示例，我们计算了成年小鼠全脑和其他不同解剖区域的细胞数量，并确定了在腺相关病毒(AAV)-PHP.eB感染全脑后用mCherry转导的细胞数量。该协议提供了更高的吞吐量，可以分析大量样本(例如，每个研究100个小鼠大脑)，为下一代生物医学研究提供平台。
Tissue-clearing techniques are powerful tools for biological research and pathological diagnosis. Here, we describe advanced clear, unobstructed brain imaging cocktails and computational analysis (CUBIC) procedures that can be applied to biomedical research. This protocol enables preparation of high-transparency organs that retain fluorescent protein signals within 7-21 d by immersion in CUBIC reagents. A transparent mouse organ can then be imaged by a high-speed imaging system (>0.5 TB/h/color). In addition, to improve the understanding and simplify handling of the data, the positions of all detected cells in an organ (3-12 GB) can be extracted from a large image dataset (2.5-14 TB) within 3-12 h. As an example of how the protocol can be used, we counted the number of cells in an adult whole mouse brain and other distinct anatomical regions and determined the number of cells transduced with mCherry following whole-brain infection with adeno-associated virus (AAV)-PHP.eB. The improved throughput offered by this protocol allows analysis of numerous samples (e.g., >100 mouse brains per study), providing a platform for next-generation biomedical research.