Rapid urine screening and staging of chronic kidney disease via NAD(P)H-activated dual-salt fluorescent probe

Chronic kidney disease (CKD) requires fast, noninvasive tools for early detection and staging that go beyond conventional serum creatinine/eGFR measurements. NAD(P)H, a key coenzyme in cellular metabolism, energy transfer, and redox signaling, is closely associated with the normal kidney function. Monitoring intracellular NAD(P)H levels offers a promising strategy for CKD assessment. Here, we present a novel dual-salt fluorescent probe (CKD-pro) for the rapid and sensitive detection of NAD(P)H. The CKD-pro is constructed from 1-methylquinolinium and naphthalene thiazolium salt units connected through vinyl linkers. Upon reduction of the quinolinium moiety by NAD(P)H, the probe converts into a donor-π-acceptor (D-π-A) architecture, resulting in a rapid fluorescence signal at 585 nm. Probe exhibits excellent analytical performance, with a linear response over 0-2 μM, a limit of detection (LOD) of 20 nM, and a response time shorter than 2 min. It also shows high selectivity and good biocompatibility, enabling real-time monitoring of NAD(P)H dynamics in G6PD-knockout cancer cells and HK-2 renal cells under hypoxia. In a clinical proof-of-concept study (midstream urine, n = 30 CKD patients and n = 10 healthy controls), the fluorescence signal from CKD-pro was significantly higher in CKD patients, showed an inverse correlation with eGFR, and successfully discriminated CKD from healthy individuals (AUC = 0.847). Moreover, the probe effectively stratified mild-to-moderate (stages 1-3) and severe (stages 4-5) CKD with an AUC of 0.915. Together, these findings establish CKD-pro as the first urine-based optical NAD(P)H probe for non-invasive CKD screening and staging, underscoring its clinical translation potential.