Caged luciferin for the detection of albuminuria based on the pseudo-esterase/luciferase activities of albumin

Abstract High levels of the urinary excretion of human serum albumin (HSA), known as albuminuria, are a crucial biomarker for kidney diseases and diabetes. In this study, we have developed a biomolecule-catalyzing chemiluminescence (BCL)-based assay for the rapid and sensitive detection of albuminuria using a caged luciferin. The caged luciferin was developed by introducing an ester protecting group at the C-3 carbonyl group of HuLumino1, a luciferin that reacts specifically with HSA. This modification effectively suppresses auto-oxidation of luciferin by dissolved oxygen in solution, which generates undesirable background signals. It also allows a 2-step pseudo-enzymatic reaction, whereby HSA first catalyzes the hydrolysis of the ester group in the luciferin, followed by an oxidative luminescence reaction. Utilizing the luminescence signals improved the detection sensitivity for HSA, achieving a limit of detection (LOD) of 0.12 µg/mL, which is significantly lower than that of unmodified HuLumino1 (6.63 µg/mL). Furthermore, the BCL-based assay provided comparable accuracy to ELISA in the quantification of albumin in urine samples while reducing the measurement time to 1 min per sample. The assay covers the diagnostic range of albuminuria (30 to 300 µg/mL), making it a promising tool for the early diagnosis and the routine monitoring of kidney diseases and diabetes.