A near-infrared fluorescent probe for endogenous hydrogen peroxide real-time imaging in living cells and zebrafish

Hydrogen peroxide (H2O2) can be produced in mitochondria and plays a significant role in physiological metabolism. Overproduction of H2O2 is a hallmark of many diseases. Therefore, it is very important to develop a highly sensitive method for detecting H2O2 both in vivo and in vitro. Previously reported benzil-based fluorescence probes are superior to those based on boronate ester in terms of reaction selectivity. However, the near infrared (NIR) probe with biocompatibility has been rarely reported for the detection of endogenous hydrogen peroxide and the real-time imaging in biological system. Hemicyanine skeleton has been proven to be effective scaffold for NM fluorescent probes for non-invasive optical imaging in vivo. In this paper, a Cy-H2O2 probe for real-time monitoring hydrogen peroxide in organisms was designed by modifying the NIR hemicyanine framework with benzil moiety. The results showed that Cy-H2O2 exhibits high specificity and sensitivity, and has good water solubility and short response time (within 10 min) for detection of hydrogen peroxide in vitro and in vivo. The reaction mechanism was deduced by detecting product of the fluorescent probe reacting with hydrogen peroxide using HPLC. The probe shows a good linear relationship for the specific response to H2O2 within the concentration range of 0-7 mu M and the detection limit is 65 nM. In addition, the probe Cy-H2O2 has been successfully applied to H2O2 detection in living cells and zebrafish.