ABSTRACT Early detection of pancreatic disease requires non‐invasive, highly sensitive biomarker detection. Here, we report a fluorescence‐based method for rapid, ultrasensitive detection of urinary trypsin without sample pretreatment. A π‐extended pyrene fluorescent probe functionalized with four carboxylate groups was designed to facilitate selective interaction and micellar aggregation with protamine, a trypsin‐cleavable substrate. Protamine‐induced aggregation results in amplified fluorescence quenching, which is efficiently reversed by trypsin‐mediated proteolysis. Trypsin cleavage reduces the arginine‐rich positive charge density and length of protamine, disassembling the compound 1 ‐protamine aggregates and recovering monomeric fluorescence, enabling highly sensitive detection. The sensing platform achieved a limit of detection (LOD) of 2.0 ng/mL in 10% human urine, highlighting its strong analytical performance. The developed sensor exhibited excellent selectivity and enabled reliable quantification of trypsin at sub‐microgram levels (0.03–0.7 µg/mL) in 10% diluted human urine, making it suitable for early‐stage pancreatic disease screening. Its clinical applicability was further validated in the presence of acute pancreatitis (AP)‐inducing factors, including aspirin, ibuprofen, dexibuprofen, acetaminophen, 6‐mercaptopurine, as well as ethanol and its metabolites (acetaldehyde and acetate). In addition, urine samples were successfully analyzed using a smartphone‐integrated fluorescence detection system, highlighting its practicality and clinical potential for early pancreatic disease diagnosis.
Han et al. (Thu,) studied this question.