An improved method to measure nitrate/nitrite with an NO-selective electrochemical sensor

Area of Science:

• Biochemistry
• Cell Biology
• Analytical Chemistry

Background:

• Nitric oxide (NO) is a vital cell signaling molecule.
• Measuring low NO levels, particularly from endothelial NO synthase (eNOS), is challenging.
• Accurate quantification of NO is essential for understanding physiological and pathophysiological processes.

Purpose of the Study:

• To develop a highly sensitive and convenient analytical method for measuring nitric oxide (NO) levels.
• To enable accurate quantification of low nanomolar concentrations of NO(x) (nitrite plus nitrate) in cell culture media.
• To provide a reliable tool for studying NO release from various cell types, including those with low basal NO production.

Main Methods:

• Utilized an ultra-sensitive NO-selective electrochemical sensor (AmiNO700).
• Implemented a nitrate reductase system coupled with glucose-6-phosphate dehydrogenase to convert nitrate to nitrite.
• Employed an acidic iodide bath for the reduction of nitrite to NO for electrochemical detection.

Main Results:

• The method accurately measures NO(x) concentrations as low as 1 to 2 nM.
• Successfully quantified basal and stimulated NO release from cultured endothelial cells.
• Demonstrated the reliability and convenience of the developed analytical assay.

Conclusions:

• The described analytical method offers high sensitivity and convenience for NO measurement.
• This assay is particularly valuable for quantifying low, physiologically relevant NO levels in diverse cell types.
• The improved method facilitates a better understanding of NO's role in biological systems.