Glutathione (GSH) is a major endogenous antioxidant that has a central role in cellular defense against toxins and free radicals. This protocol describes the preparation of CPDSA, a cyanine-based near-infrared (NIR) fluorescent probe for the detection of GSH in cells and in vivo. CPDSA is prepared with high yield through a simple two-step process. The first step is to react commercially available IR-780 iodide with excess anhydrous piperazine in anhydrous N,N-dimethyl formamide at 85 °C to form cyanine-piperazine (CP). The second step is the sulfonylation of CP with dansyl chloride in anhydrous dichloromethane. CPDSA selectively detects GSH in cells, and it has been shown to not react with other biothiols such as cysteine (Cys) and homocysteine (Hcy). This probe can also be used to monitor the GSH level of mouse bone marrow-derived neutrophils (BMDNs). The preparation of probe CPDSA takes 2 d, and experiments in cells and mice take 12-13 d.
Bibliographical noteFunding Information:
acknoWleDGMents This research was supported by a grant from the National Creative Research Initiative programs of the National Research Foundation of Korea (NRF) funded by the Korean government (grant no. 2012R1A3A2048814), by the Basic Science Research Program through the NRF funded by the Ministry of Education (grant no. 2013R1A1A2008511), and by the Yonsei University College of Medicine (grant no. 6-2014-0135 to J.-H.R.) and the National Natural Science Foundation of China (grant no. 21402057).