A new sensitive chemiluminescence probe, L-012, for measuring the production of superoxide anion by cells.
A sensitive chemiluminescence method for measuring the production of superoxide anion (O2-) by activated EoL-1 cells (human eosinophilic leukemia cell line) is described. Recently, we succeeded in synthesizing a new chemiluminescence probe, 8-amino-5-chloro-7-phenylpyrido[3,4-d]pyridazine-1,4(2H,3H)dione (L-012). In the presence of L-012, activated EoL-1 cells which produce reactive oxygen species generated a marked chemiluminescence with negligible background. The L-012-dependent chemiluminescence was completely abolished by 100-300 U/ml superoxide dismutase, indicating that the main reactive oxygen species detected in this reaction was O2-. The light intensity and the sensitivity of L-012 to O2- were higher than those of other chemiluminescence probes such as luminol and Cypridina luciferin analog (MCLA). Thus, L-012 would provide an improved chemiluminescence method for measuring O2- from cells.
Nishinaka
Y
Y
Pharmaceutical Research Division, Takeda Chemical Industries Ltd., Osaka, Japan.
Aramaki
Y
Y
Yoshida
H
H
Masuya
H
H
Sugawara
T
T
Ichimori
Y
Y
D015936:Complement C5a / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles; D007202:Indicators and Reagents; D007371:Interferon-gamma / Q000494:pharmacology; D007700:Kinetics; D015472:Leukemia, Eosinophilic, Acute; D008163:Luminescent Measurements; D008165:Luminol / Q000031:analogs & derivatives*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D010972:Platelet Activating Factor / Q000494:pharmacology; D011719:Pyrazines; D013482:Superoxide Dismutase / Q000494:pharmacology; D013481:Superoxides / Q000032:analysis / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D007093:Imidazoles; D007202:Indicators and Reagents; D010972:Platelet Activating Factor; D011719:Pyrazines; D013481:Superoxides; C063412:2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo(1,2-alpha)pyrazin-3-one; C081614:L 012; D009240:N-Formylmethionine Leucyl-Phenylalanine; D008165:Luminol; D015936:Complement C5a; D007371:Interferon-gamma; D013482:Superoxide Dismutase; D013755:Tetradecanoylphorbol Acetate