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Electrochemical sensors have emerged as a feasible approach for biomarker detection owing to their high sensitivity, operational simplicity, and cost‐effectiveness. However, the development of low‐cost, convenient, and common substrate materials with high effectiveness remains a significant challenge. This work presents a new kind of hybrid GO/AgNPs nanostructure (Ag@HA‐GO) for electrochemical detection of biotin‐streptavidin (SA) interactions in biomarkers. Specifically, GO was firstly modified with hexylamine (HA) to expand the interlayer spacing, and then silver nanoparticles (AgNPs) were incorporated within HA‐GO sheets, affording Ag@HA‐GO with good dispersibility and excellent electrical conductivity. To utilize it as a substrate material for detecting biotin‐streptavidin (biotin–SA) interactions, biotin was covalently attached onto the edges of GO sheets within Ag@HA‐GO. In subsequent electrochemical detection of biotin‐SA interaction, it was informed that GCE/biotin‐Ag@HA‐GO could serve as a promising substrate for a sensitive and stable electrochemical sensor, on account of systematical cyclic voltammetry curves. Thus, the designed GO/AgNPs hybrid nanocomposite is expected to act as an effective platform for stabilizing AgNPs, while Ag@GO‐HA demonstrates good stability, selectivity, and reproducibility in the electrochemical detection of biotin–SA under physiological conditions.
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