Zwitterionic Polymer Brushes Inside Nanoporous Gold Electrodes Enable Fouling-Resistant Electrochemical Biosensing

Surface-grafted polymer brushes offer a powerful way to engineer nanobio interfaces, yet threading these through the maze-like morphology of nanoporous gold (npAu) electrodes remains difficult: the extreme curvatures, narrow necks, and long diffusion paths hinder monomer transport, limiting chain growth. In this work, we systematically examine how pore topology, grafting strategies, and reaction parameters govern brush formation inside npAu and establish polymer-coated electrochemical biosensors. Electroanalytical diagnostics, cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), reveal that graft-to strategies generate sparse coverage, while graft-from strategies using surface-initiated activators regenerated by electron transfer atom transfer radical polymerization (SI-ARGET ATRP) of poly(2-hydroxyethyl methacrylate) (PHEMA) or poly(sulfobetaine methacrylate) (PSBMA) produce dense brushes, fully infiltrating the three-dimensional nanoporous network. Increasing initiator coverage or extending polymerization time progressively attenuates charge transfer. However, intermediate conditions yield the optimal balance, creating robust brush networks while preserving electrochemical activity and biosensing performance. Notably, complete passivation occurs far more rapidly on planar gold (pAu) than on thermally annealed npAu, highlighting the unique role of hierarchical porosity in regulating polymer growth. Finally, utilizing a PHEMA-coated npAu electrode as the sensing substrate of an electrochemical aptamer-based (EAB) biosensor retains pharmacologically relevant sensitivity, indistinguishable from that of unmodified electrodes, validating this approach for next-generation, fouling-resistant electrochemical biosensors.