Influence of Aptamer Surface Coverage on Small Target Recognition: A SPR and QCM‑D Comparative Study

Aptamers have emerged as promising biorecognition elements for the development of biosensors. The present work focused on the direct detection, by surface plasmon resonance (SPR) and quartz crystal microbalance with dissipation monitoring (QCM-D), of a low-molecular weight (LMW) compound (less than 200 Da) with an aptamer receptor presented as an oriented monolayer on the surface. These techniques are powerful for label-free, real-time characterization and quantification of molecular interactions at interfaces. Herein, we analyzed the influence of aptamer surface density on the recognition properties. A decrease of the surface concentration was shown to improve the affinity for the target because of a higher kinetic association constant that could be explained by a limitation of the steric hindrance of the aptamer on the surface. An aptamer folding is produced upon recognition of the LMW target that gives rise to the modification of the layer on the surface. This induces a displacement of water acoustically coupled to the sensing layer, thickness layer variation, and deviation of the refractive index increment (RII) of the target/aptamer complex from the sum of the RII of individual entities. We also demonstrated that the recognition signal was still detectable for low aptamer density (lower than 1 pmol·cm–2)