Dual-signal electrochemical immunoassay based on PAMAM@MXene-modified electrode for simultaneous detection of S100B and NSE

Elevated levels of S100 calcium-binding protein B (S100B) and neuron-specific enolase (NSE) are frequently observed in patients with acute cerebral infarction (ACI). Their combined quantification holds significant promise for early and accurate diagnosis. Here, we report a dual-signal electrochemical immunoarray based on poly(amidoamine)-grafted MXene (PAMAM@MXene) for the simultaneous detection of S100B and NSE. Dendritic PAMAM, rich in terminal amines, was covalently grafted onto conductive MXene, enhancing immunomolecule immobilization and electron transfer. The platform integrates two orthogonal signal readout strategies: square wave voltammetry (SWV) and chronoamperometry (I–t). Redox-active CS@MNPs–MB nanocomposites serve as SWV signal probes for S100B, while PdPtCu nanoalloys catalyze H₂O₂ decomposition to generate I–t signals for NSE. The two channels operate independently without mutual interference, enabling accurate, parallel quantification of both biomarkers. Both immunosensors exhibited excellent linearity from 1.0 pg/mL to 100 ng/mL, with high reproducibility, long-term stability, and selectivity. The limits of detection (LODs) were 3.4 × 10⁻³ pg/mL for S100B and 4.5 × 10⁻³ pg/mL for NSE. When applied to spiked artificial serum, the results strongly correlated with ELISA kits (R² > 0.997), and similar agreement was achieved in real clinical serum samples compared with the clinical gold standard. This dual-signal platform offers a modular and adaptable approach for multiplexed biomarker analysis in diverse clinical settings.