上海交通大学学报（医学版）

›› 2013, Vol. 33 ›› Issue (1): 121-.doi: 10.3969/j.issn.1674-8115.2013.01.025

• 技术与方法 • 上一篇    下一篇

建立基于超薄切片的高分辨成像技术解析组织的精细空间结构

吕春梅1, 孟 军1, 李晓林1, 李小卫1, 郭 妍1,2   

  1. 1.上海交通大学 系统生物医学研究院系统生物医学教育部重点实验室, 上海 200240； 2.上海交通大学 |肿瘤研究所癌基因及相关基因国家重点实验室, 上海 200032
  • 出版日期:2013-01-28 发布日期:2013-02-06
  • 通讯作者: 郭 妍, 电子信箱: yanguo@sjtu.edu.cn。
  • 作者简介:吕春梅（1985—）, 女, 硕士生；电子信箱: lvxinne@163.com。
  • 基金资助:

    国家自然科学基金（30900271, 60907044）；国家自然科学重大研究计划（91229108）

Tissue precise spatial structure determination with ultra-thin cutting high-resolution imaging technology

LÜ|Chun-Mei1, MENG Jun1, LI Xiao-lin1, LI Xiao-wei1, GUO Yan1,2   

  1. 1.Key Laboratory of the Ministry of Education for Systems Biomedicine, Institute for Systems Biomedicine, Shanghai Jiaotong University, Shanghai 200240, China； 2.National Laboratory for Oncogenes and Related Genes, Shanghai Cancer Institute, Shanghai Jiaotong University, Shanghai 200032, China
  • Online:2013-01-28 Published:2013-02-06
  • Supported by:

    National Natural Science Foundation of China, 30900271, 60907044； National Natural Science Major Research Program, 91229108

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摘要：

目的 建立基于超薄切片方法的高分辨三维成像技术，提高Z轴分辨率，并用于组织的精细结构研究。方法 组织样品用LRWhite树脂渗透聚合后，用钻石刀将组织切片切成50~1 000 nm的连续半薄或超薄切片，具体切片的厚度即Z轴的分辨率。切片经甲基蓝-洋红染色或多重抗体标记后，利用全自动显微镜对连续切片进行成像，通过二维拼接和三维重构获得组织高分辨的三维空间信息。结果 对人正常结肠组织和小鼠肝脏进行高分辨成像分析，获得比传统显微镜更高的分辨效果和更丰富的结构细节。结合多重抗体染色方法，对小鼠脑组织微管蛋白的分布进行了高分辨三维成像。结论 所建立的高分辨三维成像技术适用于各种组织的空间精细结构研究，超薄切片方法同时为超高分辨显微成像提供Z轴的解决方案。

关键词: 超薄切片, 组织, 高分辨, 显微镜, 抗体标记

Abstract:

Objective To establish a high-resolution three-dimensional imaging system with the Z-axis resolution improvement by ultra-thin cutting technology for tissue precise structure analysis. Methods Tissue samples were embedded with LRWhite resin and polymerized, which were cut into continuous slices of 50 nm-1 000 nm with diamond knife. Thereby the Z-axis resolution of 50 nm-1 000 nm was obtained. The semi-thin cutting or ultra-thin cutting slices were stained with methylation blue and basic fuchsin or labeled with antibody for structure illustration by automatic microscope. The high-resolution three-dimensional tissue spatial information was obtained with two-dimensional stitching and three-dimensional reconstruction. Results Using the established highresolution microscopic imaging technology, the fine structure of human normal colon tissues and mouse liver were observed. In combination with staining and antibody labeling technology, the distribution of tubulin in mouse brain tissues was observed by high-resolution three-dimensional imaging. Conclusion The established high-resolution three-dimensional imaging technology is applicable to the precise spatial structure of all kinds of tissues, and provides a solution to the Z-axis resolution for ultra-high-resolution imaging technology.

Key words: ultrathin cutting, tissue, high resolution, microscopy, antibody labeling