A fully automated paper-based smartphone-assisted microfluidic chemiluminescence sample to result immunoassay platform

Background: In recent years, adapting immunoassay technologies for point-of-care testing (POCT) to enable low-cost, on-site diagnostics has gained great attention. Among the various approaches to achieving this goal, microfluidic paper-based analytical devices (μPADs) have emerged as a promising platform. These devices are particularly appealing due to their low cost, simple procedures, and minimal sample consumption. Among the detection methods used in μPADs, chemiluminescence (CL) has attracted widespread attention for its high sensitivity. However, existing CL immunoassays often require expensive detection equipment and extensive manual operation, which limits their use in resource-poor areas or where trained personnel are lacking. Results: In this study, we developed a newly designed μPAD for CL immunoassays, leveraging smartphone-based image capture, valve-regulated reaction process, and app-driven result analysis to enable fully automated detection. Each element of the device was highly integrated and designed for programmable operation, enhancing reliability and stability. An app was developed to manage the entire process, maximizing the convenience of smartphones. The device's effectiveness was calibrated using rabbit IgG as a model, achieving a limit of detection (LOD) of 62.4 pg/mL, which represents an improvement compared to the colorimetric ELISA method with a LOD of 3 ng/mL (20pM). We then applied the device to detect tau protein, a biomarker associated with Alzheimer's disease and achieved a detection limit of 26.1 pg/mL, which is lower than the clinical cut-off value. Significance: Our device achieves complete automation and integrates the versatility of μPADs with the precision of CL immunoassays, offering an effective and accessible solution for POCT. By incorporating a user-friendly smartphone interface, it simplifies operation while maintain detection sensitivity, providing a practical and valuable improvement in the development of POCT technologies.