Nanogel-Modified SPR Sensors Enable Specific Biomarker Detection in Complex Media through Minimum Response Analysis

Direct, label-free quantification of biomarkers in complex biological media such as human serum remains challenging for surface plasmon resonance (SPR) due to nonspecific adsorption and matrix-induced artifacts. Here, we present an NTA-conjugated pNIPAM-co-AAc nanogel-modified SPR interface (NTA-NG) that enables reliable detection of interleukin-6 (IL-6) directly in human serum by leveraging multivalent protein binding (MPB)-driven signal amplification. Using the IL-6/IL-6R/GP130 system, His-tagged GP130 immobilized on the nanogel surface forms an IL-6-IL-6R-mediated ternary complex, enabling MPB-coupled SPR signal transduction. Compared with planar NTA-COOH surfaces and three-dimensional dextran-based CM5 interfaces, the NTA-NG platform provides enhanced robustness, signal stability, and sensitivity under complex biological conditions. We further introduce the minimum response (Rmin) as a robust analytical parameter that enhances signal discrimination and outperforms the conventional maximum response (Rmax) under matrix-rich conditions. In clinical serum samples from leukemia and multiple myeloma patients, Rmin-based analysis achieved superior diagnostic performance (AUC = 0.918) compared with Rmax (AUC = 0.833) and correlated well with ELISA.