Butyrylcholinesterase-Activated Bioluminescent Probe for the Ultrasensitive Whole Blood Diagnosis of Liver Diseases

Butyrylcholinesterase (BChE) is a key serum biomarker for evaluating hepatic function. However, conventional detection methods often suffer from insufficient sensitivity and strong autofluorescence interference in whole blood samples, limiting their application in early diagnosis of liver diseases. To address this issue, we herein report a strategy to develop ultrasensitive BChE-activated bioluminescent probes for detecting BChE in whole blood specimens and in vivo. Through optimization of the self-immolative linker, we identified BChE-Cl with a Cl-modified spacer as the optimal probe. BChE-Cl exhibited excellent sensitivity and selectivity toward BChE, achieving a lower limit of detection (LOD) of 4.42 U/L in whole blood. Furthermore, this optimized probe enabled sensitive visualization of endogenous BChE activity both in vitro and in vivo with significantly enhanced luminescent signals. Notably, BChE-Cl allowed reliable monitoring of BChE fluctuations in whole blood from orthotopic liver tumor and diabetes-induced liver injury mouse models, identifying a ∼2-fold increase in BChE levels in the early phase of these liver diseases. This study demonstrates that BChE-Cl is promising as a robust diagnostic tool for whole blood testing.