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Interference derived from nonspecific attachment of foulants (e.g. proteins) from food matrices (fouling) on sensor surfaces has always been a major issue in food safety sensing, deteriorating the stability and accuracy of detection. Engineering and introducing antifouling materials on sensors to strongly hydrate their surfaces is a promising approach to address this issue. Here, inspired by a bioactive peptide with antithrombotic function, a nove l antifouling zwitterionic peptide (Z-peptide) with a sequence of NH 2 -CPPPPEKDQDK (C′−N') was designed. The antifouling performance of the proposed Z-peptides modified sensor surface was demonstrated in different protein solutions and milk samples (signal suppression rate as low as 1.87%). Molecular dynamics (MD) simulations reveal that a large number of hydrogen bonds and strong interaction forces are formed between Z-peptides and water molecules, contributing to the formation of a dense hydration layer on the sensor surface to resist nonspecific adsorption. An electrochemical aptasensor based on the Z-peptide was developed for tetracycline (TC) detection. The proposed sensor was able to detect TC with a wide linear range (0.01–100 ng mL−1) and a low limit of detection (LOD = 2.91 pg mL−1, S/N ﹦3) and showed favorable specificity and stability. Given the excellent resistance to nonspecific adsorption, the antifouling sensor was capable of assaying TC in milk by diluting without other pretreatments (recoveries: 107.17%–110.16%). • A novel bioactive peptide-inspired antifouling zwitterionic peptide was designed. • The powerful hydrophilicity of the zwitterionic peptides was proved. • Sensor surface based on zwitterionic peptides shows a good antifouling effect. • An electrochemical aptasensor based the antifouling surface was developed. • The proposed sensor performed well in assaying TC residue in diluting milk samples. [ABSTRACT FROM AUTHOR]