1] Department of Chemistry and Nano Science, Global Top 5 Research Program, Ewha Womans University, Seoul, Korea. [2] Key Laboratory for Advanced Materials and Institute of Fine Chemicals, East China University of Science and Technology, Shanghai, People's Republic of China. [3].
1] Center for Biofunctional Molecules, Department of Chemistry, Yonsei University, Seoul, Korea. [2].
Nat Protoc. 2014;9(6):1245-54. doi: 10.1038/nprot.2014.086. Epub 2014 May 1.
The zinc ion has a key role in a variety of physiological and pathological processes. As a consequence, the development of sensitive and reliable methods to monitor the presence of zinc ions in cells and organisms is of great importance to biological research and biomedical applications. This protocol describes detailed procedures for the five-stage synthesis of a zinc ion-selective, cyanine-based fluorescent probe, CTMPA, from 2,6-bis(hydroxymethyl)pyridine. In addition, we describe its applications in the detection of Zn(2+) released during apoptosis in cells and endogenous Zn(2+) in living zebrafish. Notably, the use of CTMPA enabled our research group to monitor for the first time the presence of zinc ions in neuromasts of zebrafish via fluorescence. The approximate time frame for the synthesis of CTMPA is 4-5 d, and for its use in bioimaging is 8-10 h for cells and 2 h for zebrafish.
锌离子在多种生理和病理过程中都起着关键作用。因此，开发灵敏可靠的方法来监测细胞和生物体内锌离子的存在对于生物研究和生物医学应用非常重要。本方案描述了从 2,6-双(羟甲基)吡啶合成锌离子选择性、菁基荧光探针 CTMPA 的五阶段合成过程。此外，我们还描述了它在检测细胞凋亡过程中释放的 Zn(2+)和活体斑马鱼内源性 Zn(2+)中的应用。值得注意的是，使用 CTMPA 使我们的研究小组能够首次通过荧光监测斑马鱼感觉小体中锌离子的存在。合成 CTMPA 的大致时间框架为 4-5 天，用于细胞生物成像的时间为 8-10 小时，用于斑马鱼的时间为 2 小时。
