A nonradioactive assay for nitric oxide synthase activity in tissue extracts.
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.
Sutherland
Hazel
H
Department of Human Anatomy and Cell Biology, University of Liverpool, UK.
Zolle
Olga
O
Khundkar
Roba
R
Simpson
Alec W M
AW
Jarvis
Jonathan C
JC
Salmons
Stanley
S
D000818:Animals; D002956:Citrulline / Q000737:chemistry; D019793:Fluorescein / Q000737:chemistry*; D018482:Muscle, Skeletal / Q000201:enzymology*; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D013050:Spectrometry, Fluorescence / Q000379:methods; D014020:Tissue Extracts / Q000378:metabolism*
C111205:4,5-diaminofluorescein; D014020:Tissue Extracts; D002956:Citrulline; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D019793:Fluorescein