Messerli J M, Perriard J C
Institute for Cell Biology, Swiss Federal Institute of Technology, Zurich.
Microsc Res Tech. 1995 Apr 15;30(6):521-30. doi: 10.1002/jemt.1070300609.
Confocal light microscopy has found its place among the standard analytical tools in cell and molecular biology. When combined with techniques such as immunofluorescence or fluorescent in situ hybridization, the spatial distribution of individual biological components can be traced within cells and tissues and, under certain circumstances, even with living samples. In this article, advanced 3D visualization techniques have been applied to analyze the distribution of myofibrillar proteins in cultured adult rat cardiomyocytes. By combining confocal immunofluorescence microscopy with specially designed three-dimensional visualization, we have obtained images which are similar to those obtained with the scanning electron microscope. The subcellular distribution of proteins expressed after transfection of cDNA is monitored in the cultured heart cells. The expressed proteins are distinguished from their endogenous counterparts by the use of an epitope tagging technique. The described methods are suitable to specifically monitor the behavior of several closely related isoprotein mutants in cell or tissue preparations.
共聚焦光学显微镜已在细胞和分子生物学的标准分析工具中占据一席之地。当与免疫荧光或荧光原位杂交等技术相结合时，单个生物成分的空间分布可在细胞和组织内追踪，在某些情况下，甚至可对活样本进行追踪。在本文中，先进的三维可视化技术已被应用于分析成年大鼠培养心肌细胞中肌原纤维蛋白的分布。通过将共聚焦免疫荧光显微镜与专门设计的三维可视化相结合，我们获得了与扫描电子显微镜所获图像相似的图像。在培养的心脏细胞中监测cDNA转染后表达的蛋白质的亚细胞分布。通过使用表位标签技术，将表达的蛋白质与其内源性对应物区分开来。所描述的方法适用于在细胞或组织制剂中特异性监测几种密切相关的同型蛋白突变体的行为。
