A label-free aptasensor for the detection of tetracycline based on the luminescence of SYBR Green I

Abstract

A novel fluorescent method based on tetracycline-binding aptamers and the luminescence of SYBR Green I (SGI) was established for the sensitive and selective detection of tetracycline. Under natural conditions, the aptamers of tetracycline show the G-quadruplex spatial structures while SGI is nearly nonfluorescent in aqueous solution. After mixture with the G-quadruplex structured aptamers, SGI can recognize and intercalate into the aptamers, resulting in a strong fluorescence emission. When the target tetracycline was added into the solution, the specific recognition and high-affinity binding of aptamers with tetracycline will induce the conformational changes of aptamers from G-quadruplex structures to hairpin structures. Thereafter, SGI will be released from the aptamer molecules, leading to the fluorescence decline. The quantitative detection of tetracycline can be achieved by measuring the fluorescence change of the system. Under the optimum conditions, the linear range of tetracycline in the milk was from 5 to 25 μg/mL, and the detection limit was as low as 0.10 μg/mL. The recoveries of the spiked milk samples were in the range of 98.98%–104.67% with the relative standard deviations (RSDs) of 0.16%–0.67%, and the results were in agreement with those from HPLC. Therefore, the biosensor based on the specific recognition of aptamers and the fluorescence properties of SGI can detect the tetracycline in milk accurately, rapidly and specifically.