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We describe a fluorescence assay for nitric oxide synthase activity based on a new indicator, 4,5-diaminofluorescein (DAF-2). The method offers the advantage of being safer and more convenient than the citrulline radioassay in common use. The rapid and irreversible binding of DAF-2 to oxidized nitric oxide (NO) enables NO production to be measured in real time. The protocol is applied to the measurement of nitric oxide synthase in crude extracts of skeletal muscle.

Original publication




Journal article


Methods mol biol

Publication Date





105 - 111


Animals, Citrulline, Fluorescein, Muscle, Skeletal, Nitric Oxide, Nitric Oxide Synthase, Spectrometry, Fluorescence, Tissue Extracts