Development of an enzyme-linked immunosorbent assay using specific monoclonal antibodies against theacrine and its application

Theacrine (l,3,7,9-tetramethyluric acid), a purine alkaloid similar to caffeine in its chemical structure, is isolated from edible Camellia assamica var. kucha and has various pharmacological activities including hypnotic effects, anti-depressant effects, anti-inflammatory and analgesic effects, and a protective effect against stress-provoked liver damage. A rapid and simple assay is required to quantify theacrine in biological samples for pharmacokinetic studies in small animals. This study aimed to establish an enzyme-linked immunosorbent assay (ELISA) for theacrine quantification in blood. Herein, we successfully obtained monoclonal antibodies (MAbs) against theacrine, MAbs C11B5, and developed an ELISA method for the fast determination of theacrine in mouse blood. The range for calibration of theacrine by ELISA was 0.156-100 μg mL^-1. The half maximuminhibitory concentration (IC₅₀) value was 1.55 μg mL^-1. The ELISA method lays a good foundation for the further research.