A colorimetric DNA chimera biosensor based on bivalent split aptamers and G-quadruplex DNAzyme to detect norfloxacin in seafood.

Norfloxacin (NOR) residue in animal-derived food poses a significant concern due to its role in fostering bacterial resistance and potential human health risks, necessitating rapid and sensitive detection methods. In this study, a colorimetric DNA chimera biosensor was developed by integrating a bivalent split aptamer pair with G-quadruplex DNAzyme for highly sensitive NOR detection. Molecular docking studies were performed to identify the key binding regions, facilitating aptamer truncation and splitting. The bivalent strategy markedly enhanced binding affinity. The presence of NOR guides the self-assembly of the functional DNA chimera, activating the G-quadruplex/hemin DNAzyme to catalyze ABTS oxidation and generate colorimetric signals. The biosensor demonstrated a linear range of 3.75-960 nM and a limit of detection of 2.97 nM, with excellent specificity. When applied to real seafood samples, it showed good accuracy and repeatability with recovery rates of 87.87-112.59%, offering a reliable platform for rapid detection of fluoroquinolone residues in seafood.