Bioluminescent\nProbes for the Detection of Superoxide\nand Nitric Oxide

The regulation of reactive oxygen\nspecies (ROS) such\nas superoxide\n(SO) and nitric oxide (NO) is crucial in biology, influencing metabolism\nand signaling pathways. Imbalances in these species lead to oxidative\nstress and various diseases. Traditional methods for measuring SO\nand NO face challenges in terms of sensitivity and specificity, particularly\nin complex biological matrices. This report introduces bioluminescent\nprobes that leverage the intrinsic sensitivity of bioluminescence\nfor direct and selective detection of SO and NO. These probes release\nanalogs of d-luciferin upon reaction with their target ROS.\nFollowing addition of luciferase, luminescence is generated proportional\nto the amount of accumulated luciferin, allowing for quantitation\nof SO or NO. Both probes exhibit high specificity, confirmed through\ncell-free assays and cell-based studies in macrophages, demonstrating\ntheir utility in measuring cellular SO and NO production. These assays\noffer a robust, high-throughput platform for studying ROS, providing\ndirect insights into oxidative stress-related mechanisms.