Stochastic Time‐Resolved Opto‐Electrochemical Analysis of Free Nanourchin‐Conjugated Antibody Interactions With Breast Cancer Biomarkers in Serum

The combined opto-electrochemical (Opto-EC) platform enables non-invasive, real-time monitoring of biochemical kinetics, electron-transfer processes, and surface-binding events. Here, we detect the CA15-3 breast-cancer biomarker using free and randomly gold-nanourchin (GNU)-conjugated antibodies (mAb) within a serum droplet, integrated into a time-resolved Opto-EC immunoassay on an indium-tin-oxide (ITO) film. Biolayer formation on the ITO surface alters charge-transfer dynamics and reduces surface-potential differences. Time-resolved FT-NIR spectroscopy reveals peaks at 4083, 4555, and 5560-6000 cm^-1, associated with overtone and combination bands of OH, NH, and mixed NH/CH vibrations. Stochastic FT-NIR and SERS fluctuations reflect time-dependent conformational changes, binding-site accessibility, and vibrational-mode variations. Correlation-coefficient dynamics indicate shifts in the analyte's molecular environment, influencing PCA clustering. A time-mapped Raman heatmap further tracks evolving peak positions and intensities, offering complementary insight into biomarker-sensor interactions. The free and randomly conjugated antigen-antibody system exhibits nonlinear stochastic behavior.