A colorimetric aptasensor for chloramphenicol in fish based on double-stranded DNA antibody labeled enzyme-linked polymer nanotracers for signal amplification

A sensitive and facile colorimetric aptasensor for chloramphenicol (CAP) detection based on enzyme-linked polymer nanotracers labeled by double-stranded DNA (ds-DNA) antibody. In this work, the ds-DNA containing thiolated aptamer for CAP (Apt) and its complementary strand (cDNA) was immobilized on Fe3O4@Au magnetic nanoparticles (AuMNPs) as capture probe. An enzyme-linked polymer nanotracer was fabricated by co-immobilizing the ds-DNA antibody and horseradish peroxidase (HRP) labeled nanogold on Envision reagents (ds-DNA Ab/EV-AuNPs-HRP) as signal tag. EnVision reagent (EV) contains about 100 HRP and some anti-IgG antibody molecules. In the presence of CAP, the aptamer preferentially bound with CAP to release ss-DNA that cannot be recognized by ds-DNA Ab in the nanotracers and then the signal tag was substituted into supernatant after magnetic separation. The supernatant containing numerous HRP could efficiently catalyze 3,3',5,5'-tetramethylbenzidine (TMB)-H2O2 solution for the color development quantified by ultraviolet-visible spectroscopy. Experimental results showed the CAP detection owning a linear range of 0.05-100 ngmL(-1) and sensitively respond down to 0.015 ng mL(-1). Besides, the results of this method for CAP detection in the fish samples agreed well with the ELISA results, verified its accuracy and reliability. The developed method can be used for in situ CAP detection in food safety. (C) 2015 Elsevier B.V. All rights reserved.