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Several fluorescent probes were designed for the measurement of nitric oxide (NO), however, questions arose regarding their specificity and sensitivity in biological samples. In the present study we tested the reaction of a novel rhodamine-based chromophore diaminorhodamine-4M (DAR-4M) with NO and other reactive nitrogen and oxygen species.

We performed fluorometry in 96-well plates in a cell-free buffer with similar ion concentrations as the cytoplasm. Dose–response curves were generated using various NO donors and reactive nitrogen and oxygen species. The effects were compared between the red-fluorescent DAR-4M and its green-fluorescent counterpart 4-amino-5-methylamino-2′,7′-difluorofluorescein (DAF-FM).

DAR-4M had a markedly higher fluorescence yield to NO donors than DAF-FM, while both probes had a comparable threshold of sensitivity (in …