Tissue libraries enable rapid determination of conditions that preserve antibody labeling in cleared mouse and human tissue.
Tissue libraries enable rapid determination of conditions that preserve antibody labeling in cleared mouse and human tissue.
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DOI：
10.7554/elife.84112
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发表时间：
2023-01-19
期刊：
影响因子：
7.7
通讯作者：
Cardona, Albert
中科院分区：
文献类型：
--
作者：
Zwang, Theodore J.;Bennett, Rachel E.;Lysandrou, Maria;Woost, Benjamin;Zhang, Anqi;Lieber, Charles M.;Richardson, Douglas S.;Hyman, Bradley T.;Cardona, Albert
难以实现完整、特异和均匀的染色是阻碍组织透明技术广泛应用于对大量人体组织进行成像的主要瓶颈。在这份手稿中，我们描述了一种快速设计免疫染色方案的程序，用于清除脑组织的抗体标记。我们准备了0.5-1.0 mm厚的组织切片文库，通过类似但不同的程序进行固定、预处理和清除，以优化一组抗体的染色条件。来自小鼠组织库的结果与类似准备的人脑组织库的结果很好地相关，表明小鼠组织是方案优化的适当替代品。这些数据表明，程序上的差异并不会以相同的方式影响每个抗体-抗原对，微小的变化可能会产生有害的影响，因此，应该针对每个目标进行优化。这里概述的方法将有助于指导研究人员成功标记各种目标，从而消除获取大量、清晰的人体组织中丰富的3D信息的主要障碍。
Difficulty achieving complete, specific, and homogenous staining is a major bottleneck preventing the widespread use of tissue clearing techniques to image large volumes of human tissue. In this manuscript, we describe a procedure to rapidly design immunostaining protocols for antibody labeling of cleared brain tissue. We prepared libraries of 0.5–1.0 mm thick tissue sections that are fixed, pre-treated, and cleared via similar, but different procedures to optimize staining conditions for a panel of antibodies. Results from a library of mouse tissue correlate well with results from a similarly prepared library of human brain tissue, suggesting mouse tissue is an adequate substitute for protocol optimization. These data show that procedural differences do not influence every antibody-antigen pair in the same way, and minor changes can have deleterious effects, therefore, optimization should be conducted for each target. The approach outlined here will help guide researchers to successfully label a variety of targets, thus removing a major hurdle to accessing the rich 3D information available in large, cleared human tissue volumes.