Biophotonic Sensor for Real-time and Non-invasive Detection of Extracellular H2O2 Released by Stimulated Cells

A highly sensitive biophotonic tool was developed that performs real-time detection of scarce H2O2 concentrations down to the subnanomolar range. The detection method is based on the dark-field spectroscopic analysis of cytochrome c (cytc c) embedded into a matrix (polystyrene beads, porous membranes) that exhibits a high scattering cross-section. Therefore, the incoming light undergoes multiscattering(1) in the matrix sheltering cyt c - i.e. an increase of the optical trajectory-which leads to an improved signal-to-background ratio of the cyt c spectrum recorded. Then, the analysis of the absorption peak at 550 nm and its conversion into a normalized redox state coefficient phi provides a sensitive indication of cyt c oxidation state and consequently its oxidation rate in the presence of H2O2. A limit-of-detection at the subnanomolar range was achieved using this H2O2 sensing principle. Moreover the device was able to sense the dynamics of H2O2 extracellular release by human cells exposed to oxidative-stress inducers.