Table 1.
Summary of methods for the detection of nitric oxide–derived oxidants
| Method | NO• | NO2• | N2O3 | NO2− | NO3− | DNICs | RSNO | HNO | ONOO−/ONOOH | CO3•− | HO• |
|---|---|---|---|---|---|---|---|---|---|---|---|
| Oxyhemoglobin oxidation, UV-visible, <1 μm | Yes, fast | Yes, but very slow | Yes, slower than NO• | ||||||||
| NO•-selective electrode | Yes, in neutral buffer | No, but NO2− forms NO• in acid | No, only with Cu(II) and cysteine | ||||||||
| EPR with spin traps | Yes, cPTIO (forms NO2•) or Fe(MGD)2 | Yes, nitromethane and nitrone traps | Yes, DMPO and other traps | ||||||||
| Direct EPR, 200 nm sensitivity | Nitrosyl heme with Fe(II) heme, 77 K | Yes | Nitrosyl heme with Fe(III) heme, 77 K | Fast flow | |||||||
| Fluorogenic diamino probes (DAN and DAF) | Not direct, but through N2O3 or oxidants and NO• | Yes, fast and direct reaction | Yes, in acidic pH | ||||||||
| NOS activity, radiolabeled arginine | Yes, controls with NOS inhibitors and without NADPH | ||||||||||
| Ozone-based chemiluminescence, high sensitivity, 10 nm | Yes, neutral buffer in purge vessel | Reductant in purge vessel | Acidic triiodide in purge vessel | Acidic triiodide in purge vessel | Acidic triiodide in purge vessel | ||||||
| Luminol chemiluminescence | Yes | Not direct but through derived free radicals | Yes | Yes | |||||||
| DCFH2 and DHR fluorescence | Yes | Not direct but through derived free radicals | Yes | Yes | |||||||
| HPLC | Yes | Yes | Yes | Low Mr RSNO | Glutathione sulfinamide | m-Tyrosine or hydroxylated probes | |||||
| UV spectrophotometry | Pulse radiolysis | Pulse radiolysis | Must be purified | Low sensitivity (50 μm) | Stocks and stopped-flow | Pulse radiolysis | |||||
| Griess for nitrite detection (micromolar sensitivity) | No, but oxidation gives NO2− | No, but hydrolysis product is NO2− | Yes | Not direct, but after reduction to NO2− | Not direct, but with Hg(II) | ||||||
| Biotin-switch proteomics | Yes, provides protein identity | ||||||||||
| N2O detection | Yes | ||||||||||
| Oxidized metalloporphyrins, UV-visible or EPR | Yes, interference by reductants and NO• | ||||||||||
| Co(III)-porphyrin electrode | Yes | ||||||||||
| Cu(II)-fluorogenic probes | Yes, interference by reductants | ||||||||||
| Nitroxide probes | Yes, interference by reductants | ||||||||||
| 2-Mercapto-2-methylpropionic acid fluorogenic probe | Yes, confirm specific product | ||||||||||
| Arylphosphine probes | Yes, RSNO interference, confirm specific product | ||||||||||
| Detection of 3-nitrotyrosine, several methods, MS confirmation desirable | Not direct, only in the presence of strong oxidants | Yes, product of NO2• with tyrosyl radical | Yes, in acid | Not direct but through derived free radicals | Forms tyrosyl radical | Forms tyrosyl radical | |||||
| Boronic acid probes | Yes, direct and fast, H2O2 reaction very slow |