Charge-Pairing Enabled Renal-Clearable Chemiluminescent Gold Nanoprobes for Ratiometric Imaging and Urinalysis of Pathological Reactive Oxygen Species

Excessive generation of reactive oxygen species (ROS) is a hallmark of many diseases and represents an important molecular signal for early disease detection. Chemiluminescent probes hold potential for ROS detection, yet they are often hindered by high mononuclear phagocyte system (MPS) uptake, limited light penetration depth, and single-signal readout <i>in vivo</i>. Herein, we report renal-clearable ratiometric chemiluminescent ROS nanoprobes (ABEI-MSA-AuNPs) by simply conjugating cationic chemiluminescent molecules, <i>N</i>-(4-aminobutyl)-<i>N</i>-ethylisoluminol (ABEI), onto the surface of anionic ultrasmall mercaptosuccinic acid (MSA)-coated gold nanoparticles (MSA-AuNPs). Strikingly, although neither ABEI nor MSA-AuNPs are intrinsically renal clearable, conjugating ABEI onto MSA-AuNPs results in efficient renal clearance and reduces the level of hepatic sequestration of AuNPs by nearly 100-fold, which is due to a charge-pairing effect that effectively suppresses opsonization and MPS recognition. The large Stokes shift of luminescent MSA-AuNPs allows energy transfer from ABEI to MSA-AuNPs for ROS-triggered chemiluminescence emission beyond 800 nm. Leveraging ROS-sensitive chemiluminescence and ROS-insensitive photoluminescence signals, we show that ABEI-MSA-AuNPs permit noninvasive detection of pathological ROS through dual-channel ratiometric imaging and ratiometric urinalysis using multiple disease models. Our discoveries highlight a charge-pairing strategy for constructing chemiluminescent probes with ultralow MPS uptake, efficient renal elimination, and ratiometric readout, showing promise in early detection of ROS-associated pathological events <i>in vivo</i>.