Detection of diabetic nephropathy biomarker cystatin C using bimetallic-based immunosensor

In this research, a new electrochemical immunosensor was created to detect Cystatin C with high sensitivity, utilizing Ag-Au bimetallic nanoparticles synthesized through eco-friendly methods. The nanoparticles, synthesized using waste tea leaves extract, exhibited uniform spherical morphology with an average diameter of 28.5 ± 4.2 nm and demonstrated excellent conductivity, increasing the electrode's effective surface area by 2.25-fold compared to the bare GCE. Cyclic voltammetry and electrochemical impedance spectroscopy confirmed successful stepwise assembly of the immunosensor, with the peak-to-peak separation (ΔEp) increasing from 111 mV at bare GCE to 136 mV after antibody immobilization and 230 mV upon antigen binding, and a charge-transfer resistance (Rct) trend consistent with interface modification. Under optimized conditions (pH 7.4, 37°C, 35-minute incubation), the immunosensor displayed dual linear ranges of 0.1–10 ng/mL and 10–100 ng/mL. The sensor achieved a remarkable limit of detection of 0.034 ng/mL and demonstrated excellent reproducibility with RSDs of 3.8 % for inter-sensor and 2.9 % for intra-assay precision. The immunosensor retained 94.3 % of its original reactivity following a 30-day storage period and exhibited excellent specificity in distinguishing against typical interferences. Validation using clinical samples revealed excellent correlation with standard ELISA (R² = 0.9943) and impressive recovery rates ranging from 96.8 % to 103.5 %. This environmentally friendly, cost-effective platform offers significant advantages in terms of analysis time (35 min versus 3.5 h for ELISA) and demonstrates great potential for point-of-care CyC detection in clinical settings.