Laser capture microdissection of mammalian tissue.
Laser capture microdissection of mammalian tissue.
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DOI：
10.3791/309
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发表时间：
2007-01-01
期刊：
影响因子：
--
通讯作者：
Edwards, Robert A
中科院分区：
文献类型：
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作者：
Edwards, Robert A
激光捕获显微镜，也称为激光显微切割 (LMD)，使用户能够从冷冻或福尔马林固定、石蜡包埋的组织切片中分离少量细胞或组织。 LMD 技术依赖于放置在组织切片顶部或下方的热不稳定膜。在一种方法中，使用聚焦激光能量将膜熔化到下面的细胞上，然后可以将其从组织切片中取出。另一方面，激光能量沿着组织上“绘制”的路径蒸发箔片，使选定的细胞落入收集装置中。每种技术都允许以几微米的最小分辨率选择细胞。 DNA、RNA、蛋白质和脂质样品可以从显微解剖样品中分离和分析。在本视频中，我们分七个部分演示了 Leica AS-LMD 激光显微切割仪器的使用，包括 LMD 原理介绍、仪器使用初始化、样品制备的一般注意事项、安装标本和设置捕获管、对准显微镜、调整捕获控制和捕获组织标本。激光捕获显微解剖使研究人员能够分离出小至几个细胞当量的纯细胞群样本。这允许分析不含邻近污染物的感兴趣细胞，这可能会混淆实验结果。
Laser capture microscopy, also known as laser microdissection (LMD), enables the user to isolate small numbers of cells or tissues from frozen or formalin-fixed, paraffin-embedded tissue sections. LMD techniques rely on a thermo labile membrane placed either on top of, or underneath, the tissue section. In one method, focused laser energy is used to melt the membrane onto the underlying cells, which can then be lifted out of the tissue section. In the other, the laser energy vaporizes the foil along a path "drawn" on the tissue, allowing the selected cells to fall into a collection device. Each technique allows the selection of cells with a minimum resolution of several microns. DNA, RNA, protein, and lipid samples may be isolated and analyzed from micro-dissected samples. In this video, we demonstrate the use of the Leica AS-LMD laser microdissection instrument in seven segments, including an introduction to the principles of LMD, initializing the instrument for use, general considerations for sample preparation, mounting the specimen and setting up capture tubes, aligning the microscope, adjusting the capture controls, and capturing tissue specimens. Laser-capture micro-dissection enables the investigator to isolate samples of pure cell populations as small as a few cell-equivalents. This allows the analysis of cells of interest that are free of neighboring contaminants, which may confound experimental results.