Preparation of a cyanine-based fluorescent probe for highly selective detection of glutathione and its use in living cells and tissues of mice.
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.
Department of Chemistry and Nano Science, Ewha Womans University, Seoul, Korea.|Key Laboratory of Pesticide and Chemical Biology, Ministry of Education, College of Chemistry, Central China Normal University, Wuhan, China.
Jun
J
Yin
Research Center for Human Natural Defense System, Yonsei University College of Medicine, Seoul, Korea.|BK 21 Plus Project for Medical Science, Yonsei University College of Medicine, Seoul, Korea.
Younghee
Y
Kwon
Department of Chemistry and Nano Science, Ewha Womans University, Seoul, Korea.
Dabin
D
Kim
Department of Chemistry and Nano Science, Ewha Womans University, Seoul, Korea.
Dayoung
D
Lee
Department of Chemistry and Nano Science, Ewha Womans University, Seoul, Korea.
Gyoungmi
G
Kim
Department of Chemistry and Nano Science, Ewha Womans University, Seoul, Korea.
Ying
Y
Hu
Research Center for Human Natural Defense System, Yonsei University College of Medicine, Seoul, Korea.|BK 21 Plus Project for Medical Science, Yonsei University College of Medicine, Seoul, Korea.
Ji-Hwan
JH
Ryu
Department of Chemistry and Nano Science, Ewha Womans University, Seoul, Korea.
Juyoung
J
Yoon
D000818:Animals; D002232:Carbocyanines / Q000378:metabolism*; D003584:Cytological Techniques / Q000379:methods*; D005456:Fluorescent Dyes / Q000138:chemical synthesis* / Q000302:isolation & purification*; D005978:Glutathione / Q000032:analysis*; D008810:Mice, Inbred C57BL
D002232:Carbocyanines; D005456:Fluorescent Dyes; D005978:Glutathione