Electrochemical Immunoassays for Point-of-Care Detection of Heart Failure Biomarkers in Saliva: Advancing Accessible Healthcare Testing

Abstract

Heart failure (HF) remains a leading cause of morbidity and mortality worldwide, with early detection and ongoing management critical for improving patient outcomes. However, current medical testing methods are often invasive, expensive, and inaccessible to many populations, particularly those in rural or resource-limited settings. Electrochemical detection offers a promising pathway to address these limitations. Electrochemical assays on screen-printed carbon electrodes (SPCEs) are an attractive option due to their low cost, favorable electrochemical properties, and ability to be fabricated in unique geometries. When combined with microfluidics, these technologies open opportunities for automated platforms, multiplexed detection, and the use of non-invasive sample matrices like saliva. However, developing a sensor compatible with saliva, engineering a microfluidic platform capable of processing viscous samples, and integrating these components into a unified system all present significant challenges. This dissertation addresses these issues by developing a novel, non-invasive electrochemical immunoassay platform for the detection of HF biomarkers in saliva, towards rapid, affordable, and decentralized point-of-care testing (POCT), and exploring simpler detection modalities. By leveraging saliva as a sample and integrating advanced sensor technologies, this work aims to bridge the gap between clinical need and technological capability, ultimately supporting more equitable healthcare delivery. Chapter 2 introduces the development of an electrochemical immunosensor specifically designed for the detection of galectin-3 (Gal-3) in saliva. The sensor utilizes SPCEs and optimized surface chemistry to ensure stability and sensitivity suitable for clinical applications. The chapter details strategies to overcome challenges associated with the complex saliva matrix, including effective antibody immobilization and sample preparation protocols, resulting in reliable biomarker quantification. Chapter 3 expands the platform to enable multiplexed detection of both Gal-3 and S100A7, two biomarkers relevant to HF prognosis. A dual-electrode array is integrated into a capillary-driven microfluidic device, allowing simultaneous detection of multiple analytes from a single saliva sample. The device automates reagent delivery and streamlines the assay workflow, requiring minimal user intervention and delivering results rapidly and cost-effectively. In the microfluidic system, the assay quantifies salivary levels of Gal-3 and S100A7, demonstrating successful multiplex detection and differentiation between these biomarkers. Chapter 4 investigates surface modification and antibody immobilization strategies for label-free immunosensors using SPCEs. Commercial SPCEs are identified as superior to lab-fabricated counterparts due to their enhanced consistency and electrochemical performance. Air-plasma treatment is shown to enhance electrode properties, and a comparison of immobilization methods-including passive adsorption, Protein A binding, and EDC/NHS coupling-provides insight into optimal strategies for sensor performance. A proof-of-concept immunoassay for HF biomarker Gal-3 validates the platform’s sensitivity and clinical relevance, advancing label-free SPCE-based biosensors for decentralized testing. This dissertation establishes a foundation for next-generation HF testing by demonstrating that non-invasive, saliva-based electrochemical immunoassays can deliver clinically relevant results at the point-of-care. The developed platform combines affordability, ease of use, and adaptability, making it suitable for widespread deployment in diverse healthcare environments. By reducing barriers to regular monitoring and early intervention, this work has the potential to transform HF management, reduce hospitalizations, and ultimately improve patient outcomes. Future directions include expanding the biomarker panel, further automating the testing process, and integrating digital health solutions to enhance remote patient monitoring and disease management.