Elucidating the dual detection mechanisms of functionalized gold nanocluster-based biosensor for quantitative...

Reliable and sensitive heparin detection in clinical settings is essential for safe anticoagulant therapy and prevention of overdose associated complications. In this study, self-assembled chitosan-functionalized gold nanoclusters (Ch-AuNCs) were synthesized, where amine-rich cationic chitosan facilitated a confinement effect that enhanced photoluminescence through aggregation induced effect (AIE). Ch-AuNCs exhibited both fluorescence and peroxidase-like catalytic activity, enabling dual-mode sensing platform for heparin detection. The biosensor demonstrated excellent analytical performance, with wide linear range of 0.25-15 U/mL for fluorimetric quenching (LOD: 0.016 U/mL) and 0-30 U/mL for colorimetric enhancement (LOD: 0.023 U/mL), covering both therapeutic and overdose concentrations. Molecular analyses elucidated the nature of interaction, binding thermodynamics, and high specificity of Ch-AuNCs with heparin over structurally similar glycosaminoglycans. Mechanistic studies revealed a dual effect of heparin on the Ch-AuNCs system that includes disassembly of the self-assembled AIE structure, and electrostatic bridging of multiple nanoclusters. The fabricated sensor was validated in FBS, human plasma and serum, showing recoveries of 93-112% and a strong correlation with the standard aPTT assay. Its practical applicability was demonstrated using paper-based smartphone platforms for user-friendly heparin detection. In summary, the study presents a highly sensitive, selective and cost-effective dual-mode biosensor with a potential for translation in clinical settings as point-of-care device.