Rapid On-Demand Point-of-Care Monitoring of Clozapine and Its Metabolite Norclozapine Using Miniature Mass...

Background/Objectives: Clozapine remains the gold standard for treatment-resistant schizophrenia. However, its narrow therapeutic window and risk of severe side effects require close monitoring of both clozapine and its primary metabolite, norclozapine. Existing therapeutic drug monitoring (TDM) methods are limited by delays, high costs, and operational complexity. This study introduces three rapid point-of-care (POC) assays utilizing a miniature mass spectrometer (Mini-MS) to quantify clozapine and norclozapine in plasma, whole blood, and dried blood spots (DBSs), facilitating applications across diverse clinical settings. Methods: The analytical performance of the assay was evaluated for sensitivity, specificity, reproducibility, and correlation with reference methods. Clinical samples from two hospitals were analysed and validated against conventional liquid chromatography tandem mass spectrometry (LC-MS/MS) reference standards at New South Wales Health Pathology (NSWHP) and Tsinghua University laboratories. Results: The Mini-MS assay accurately quantified both analytes within therapeutic ranges across all matrices. Inter-assay coefficients of variation ranged from 7.9 to 14.1% for clozapine and from 1.6 to 14.6% for norclozapine. Accuracy fell between 85 and 117% in plasma and blood extracts. Strong linearity was demonstrated (R² = 0.98-0.99) over the concentration range of 10-1000 ng/mL. Results from the Mini-MS analysis showed excellent correlations with LC-MS/MS results (r = 0.998). Conclusions: In this proof-of-concept study, the Mini-MS-based POC assays enable rapid, reliable quantification of clozapine and norclozapine, with performance comparable to conventional laboratory methods. This platform supports real-time TDM, facilitating timely dose adjustments, adherence monitoring, and ultimately improving patient outcomes.