Design and Characterization of a Nano-Encapsulated Self-Referenced Fluorescent Nitric Oxide Sensor for Wide-Field Optical Imaging

A nano-encapsulated fluorescence dye DAF-2 sensor that is specifically sensitive to nitric oxide (NO) was fabricated by using an electrostatic layer-by-layer (LbL) self-assembly technique. Fluorescence calibrations of the NO sensor were collected by a wide-field optical imaging system and a fluorescence spectrometer using NO standards generated by the self decomposition of S-nitrosol-acetyl-penicillamine (SNAP). The NO sensor consists of two fluorescence dyes, indicator DAF-2 and a reference R-PE. The two dyes share the same excitation, and have different emission wavelengths. The calibration results show that the indicator fluorescence intensity increases with NO standard solution additions, while the reference fluorescence intensity does not. The encapsulation reduced the NO sensor detection limit, but it is still within physiological level of NO. This sensor design can also achieve ratiometric self referencing with very little leaching effect.