Development of a SPR aptasensor containing oriented aptamer for direct capture and detection of tetracycline in multiple honey samples.

Although surface plasmon resonance (SPR) technique and aptamer technology shows great potential in analytical and biological chemistry, direct capture and analysis of small molecules using SPR remains tough. Detection sensitivity of aptasensor and recognition ability of aptamer is limited, because direct immobilization of aptamer causes large steric hindrance and strand entanglement. Herein, we chose a typical small molecule-tetracycline (Mw. 444.4 g/mol) as a model, and combined aptamer technology, DNA nanostructure, and commercial Biacore T200 SPR instrument to develop a straightforward format SPR aptasensor. Anti-tetracycline aptamer (Apt76) was fabricated on the top of a tetrahedron nanostructure to provide a better accessibility to tetracycline than the single-stranded Apt76 (ss-Apt76), and thus to improve sensitivity of the SPR aptasensor. The aptasensor was then validated in real world application for tetracycline screening in multiple honey samples, achieving good recovery rates of 80.20-114.3%, intuitive sensorgrams indicating the binding kinetic properties, and high specificity towards tetracycline. LOD of the tetrahedron-based SPR aptasensor was obtained using the real honey sample and calculated to be 0.0069 μg/kg, which was 10-fold range lower than that of the ss-Apt76-based aptasensor. The proof-of-concept demonstrated that aptamers of small molecules can be oriented immobilized on the SPR surface in a uniform nanoscale distance in both lateral and vertical direction, so as to achieve better conformational folding and better accessibility to small molecules. The concept is promising to be a universal and powerful tool for other ligand immobilization and SPR studies for both real world detection and molecular interaction.