Bkaily Ghassan, Al-Khoury Johny, Simon Yanick, Jacques Danielle
Department of Anatomy and Cell Biology, Faculty of Medicine, University of Sherbrooke, 2500 boul. De l'Universite, Sherbrooke, QC, Canada, JIH 5N4.
Methods Mol Biol. 2017;1527:177-187. doi: 10.1007/978-1-4939-6625-7_14.
Confocal microscopy, coupled to high-performance hardware and software systems, has provided scientists with the capability of overcoming some of the limitations of standard microscopic imaging measurements of intracellular ions. The technique for loading of ion fluorescent probes is easily achieved; however, the quality of calcium measurements depends on the way of using the confocal system. In order to optimize this technique, scientists need to be familiar with the basic approaches and limitations of confocal microscopy. In this chapter, we will describe sample preparation, fluorescent probe loading, labeling of intracellular compartments, and the setting of parameters as well as protocols for measurements and limitations of the technique.
共聚焦显微镜与高性能硬件和软件系统相结合，为科学家提供了克服细胞内离子标准显微成像测量某些局限性的能力。离子荧光探针的加载技术很容易实现；然而，钙测量的质量取决于共聚焦系统的使用方式。为了优化这项技术，科学家需要熟悉共聚焦显微镜的基本方法和局限性。在本章中，我们将描述样品制备、荧光探针加载、细胞内区室标记、参数设置以及该技术的测量方案和局限性。
