Enhanced Detection of Enrofloxacin in Seawater Using a Newly Selected Aptamer on a Graphite Oxide-Based Biosensor

Abstract

Developing aptasensors offers several advantages including sensitivity, selectivity, cost-effectiveness, and speed over traditional analytical techniques for antibiotic detection. We have successfully identified Enro_ap3, a 30-mer enrofloxacin-binding aptamer with micromolar binding affinity, through an optimized Capture-SELEX platform. Compared to other reported enrofloxacin-binding aptamers, this shorter aptamer not only streamlines the design process but also eliminates the common issue of strong nonspecific binding to the GO surface, thereby improving the overall detection capabilities of the biosensor (GO-Enro_ap3-FAM). This GO aptasensor demonstrated remarkable selectivity by effectively distinguishing enrofloxacin from different structurally diverse antibiotics. The sensor boasts a LOD of 32.15 μg/mL, 2.5 times more sensitive than the original 30-mer, with recoveries of 74%–92% and relative standard deviations of 6.3%–12.5% in seawater samples spiked with enrofloxacin. Furthermore, the GO aptasensor’s detection capabilities were found to be on par with traditional LC-MS/MS techniques, exhibiting no significant differences in recovery rates even in complex matrices. The sensor’s performance remained consistent across variations in salinity, acidity, and total organic carbon concentrations in seawater samples collected from different locations, underlining its robustness in diverse environmental conditions and its suitability for real-world seawater monitoring applications. Our findings highlight the importance of the aptamer’s chain length and its binding affinity toward the target after immobilization on the GO substrate. These factors significantly impact the performance of GO aptasensors in seawater. Overall, the GO aptasensor provides a well-balanced approach, combining sensitivity, environmental adaptability, and practical usability for detecting pharmaceutical contaminants, such as antibiotics, in marine environments.