Using Metalloporphyrin Nanosensors for In Situ Monitoring and Measurement of Nitric Oxide and Peroxynitrite in a...

Nitric oxide (NO) is an inorganic signaling molecule that plays a crucial role in the regulation of numerous physiological functions. An oxidation product of the cytoprotective NO is cytotoxic peroxynitrite (ONOO^-). In biological systems, the concentrations of NO and ONOO^- are typically transient, ranging from nanomolar to micromolar, and these increases are normally followed by a swift return to their basal levels due to their short life spans. To understand the vital physiological role of NO and ONOO^- in vitro and in vivo, sensitive and selective methods are necessary for direct and continuous NO and ONOO^- measurements in real time. Because electrochemical methods can be adjusted for selectivity, sensitivity, and biocompatibility in demanding biological environments, they are suitable for real-time monitoring of NO and ONOO^- release. Metalloporphyrin nanosensors, described here, have been designed to measure the concentration of NO and ONOO^- produced by a single human neural progenitor cell (hNPC) in real time. These nanosensors (200-300 nm in diameter) can be positioned accurately in the proximity of 4-5 ± 1 μm from an hNPC membrane. The response time of the sensors is better than a millisecond, while detection limits for NO and ONOO^- are 1 × 10^-9 and 3 × 10^-9 mol/L, respectively, with a linear concentration response of up to about 1 μM. The application of these metalloporphyrin nanosensors for the efficient measurement of the concentrations of NO and ONOO^- in hNPCs is demonstrated, providing an opportunity to observe in real time the molecular changes of the two signaling molecules in situ.