Defect Engineering of DNA Origami ROS Sensors for Portable Urinalysis

Translating in situ dynamic changes of key signaling molecules into actionable clinical readouts remains a formidable challenge for noninvasive diagnostics. Here, focusing on reactive oxygen species (ROS) as pivotal signaling mediators, we developed defect-programmed DNA origami ROS sensors (DOSs) for portable urinalysis of localized oxidative stress. Using triangular DNA origami (DO) nanostructures as two-dimensional synthetic soft crystals, we programmed the number of discontinuity defects between adjacent staple strands and established a positive correlation between defect number and ROS-triggered degradation kinetics. To transform this programmable degradation into a diagnostic function, we then engineered DOSs via orthogonal assembly of targeting and signaling modules onto DO. In a murine model of acute liver injury (ALI), DOSs selectively accumulated in the liver and underwent ROS-triggered fragmentation into renal-clearable debris, converting hepatic ROS levels into quantifiable urinary signals. Notably, this transformation efficiency depended positively on defect number in DOSs, enabling portable urinalysis that detected ALI onset at least 4 h earlier than conventional alanine aminotransferase (ALT) testing, with a maximum area under the curve of 0.94. This defect-engineering strategy establishes a generalizable platform for early, noninvasive diagnosis of ROS-related diseases.