<p>Kanamycin (KANA), a widely used aminoglycoside antibiotic, may accumulate in food products and pose potential risks to human health, calling for sensitive and reliable analytical strategies. A dual-mode detection method integrating surface-enhanced Raman spectroscopy (SERS) with colorimetric sensing was developed. This approach enabled sensitive and rapid detection of KANA. A core–shell Au@CeO₂ nanozyme-mediated aptasensor was synthesized, which exhibited oxidase-like activity and enhanced SERS performance. Without KANA, adsorption of the aptamer onto the nanozyme surface inhibited the nanozyme-catalyzed oxidation of 3,3’,5,5’-tetramethylbenzidine (TMB), leading to a simultaneous decrease in both SERS and colorimetric signals. Under optimized conditions, the aptasensor enabled quantitative KANA analysis with favorable sensitivity and selectivity, achieving limits of detection of 3.20 × 10^–10 mol/L (SERS mode) and 5.10 × 10^–6 mol/L (colorimetric mode), with linear ranges of 1.00 × 10^–9-1.00 × 10^–5 mol/ L (SERS) and 5.00 × 10^–5-5.00 × 10^–3 mol/L (colorimetric). The practicality of this platform was validated in milk and egg white samples, with recoveries ranging from 98.6% to 107.0% in milk and from 95.6% to 105.0% in egg white. In this strategy, Au@CeO₂ is designed to function simultaneously as an oxidase-like nanozyme and a SERS-active substrate. Although the two modes differ in sensitivity and linear range, they provide complementary analytical information: the SERS mode is suitable for trace-level quantitative analysis, whereas the colorimetric mode offers an auxiliary visual response at relatively higher KANA concentrations. This work provides a dual-readout approach for KANA analysis in food samples.</p> Graphical Abstract <p></p>