Selective detection of superoxide anion radicals generated from macrophages by using a novel fluorescent probe.
Quantitation of superoxide radical (O (2)(-).) production at the site of radical generation remains challenging. A simple method to detect nanomolar to micromolar levels of superoxide radical in aqueous solution has been developed and optimized. This method is based on the efficient trapping of O(2)(-). using a novel fluorescent probe (2-chloro-1,3-dibenzothiazolinecyclohexene), coupled with a spectra character-signaling increase event. A high-specificity and high-sensitivity fluorescent probe was synthesized in-house and used to image O(2)(-). in living cells. Better selectivity for O(2)(-). over competing cellular reactive oxygen species and some biological compounds illustrates the advantages of our method. Under optimal conditions, the linear calibration range for superoxide anion radicals was 5.03 x 10(-9)-3.33 x 10(-6) M. The detection limit was 1.68 x 10(-9) M. Fluorescence images of probe-stained macrophages stimulated with 4beta-phorbol 12-myristate 13-acetate were obtained successfully using a confocal laser scanning microscope.
Gao
Jing Jing
JJ
College of Chemistry, Chemical Engineering and Materials Science, Shandong Normal University, Jinan, China.
Xu
Ke Hua
KH
Tang
Bo
B
Yin
Ling Ling
LL
Yang
Gui Wen
GW
An
Li Guo
LG
D000818:Animals; D002374:Catalase / Q000737:chemistry; D002460:Cell Line; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008264:Macrophages / Q000737:chemistry* / Q000166:cytology / Q000187:drug effects; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008956:Models, Chemical; D015394:Molecular Structure; D017382:Reactive Oxygen Species / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013482:Superoxide Dismutase / Q000737:chemistry; D013481:Superoxides / Q000032:analysis* / Q000737:chemistry; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019820:Xanthine / Q000737:chemistry; D014969:Xanthine Oxidase / Q000737:chemistry
D005456:Fluorescent Dyes; D017382:Reactive Oxygen Species; D013481:Superoxides; D019820:Xanthine; D002374:Catalase; D013482:Superoxide Dismutase; D014969:Xanthine Oxidase; D013755:Tetradecanoylphorbol Acetate