A label-free colorimetric aptasensor based on an engineered chimeric aptamer and Au@FeP nanocomposites for the detection of kanamycin

The abuse of antibiotics such as kanamycin (KAN) has caused severe environmental pollution and health threat, attracting considerable attention for the monitoring of antibiotic residues. Here, a novel bivalent anti-KAN chimeric aptamer (Chi-Apt) was rationally designed through an efficient post-screening strategy that connected two homogenous anti-KAN aptamers with a poly-T linker. The engineered Chi-Apt achieved 34-fold enhanced binding affinity for KAN that was revealed by biolayer interferometry (BLI) measurement compared with the original aptamer. Meanwhile, molecular docking simulation and isothermal titration calorimetry (ITC) assay demonstrated that the engineered Chi-Apt has lower binding free energies and stronger non-covalent binding forces for KAN than the original aptamer. The engineered Chi-Apt was further evaluated by developing a label-free colorimetric aptasensor based on Au@FeP nanocomposites for KAN detection. The aptasensor exhibited a lower limit of detection (LOD) than that achieved by the colorimetric aptasensor based on the original aptamer, with a LOD of 0.98 nM and a dynamic response range of 2.4–150 nM. Moreover, good recoveries of 103.3–110.1% in tap water and 107.6–112.7% in milk samples were obtained by the proposed colorimetric aptasensor. The recoveries detected by the proposed colorimetric aptasensor in real samples were consistent with that obtained by HPLC analysis, suggesting the good practicality and accuracy of the colorimetric aptasensor. This study may provide an efficient strategy for engineering high affinity aptamers to construct sensitive biosensing platforms against other targets.