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Aptamer biosensors are one of the most powerful techniques in biosensing. Achieving the best platform to use in aptamer biosensors typically includes crucial chemical modifications that enable aptamer immobilization on the surface in the most efficient manner. These chemical modifications must be well defined. In this work we propose nanoporous anodic alumina (NAA) chemically modified with streptavidin as a platform for aptamer immobilization. The immobilization of biotinylated thrombin binding aptamer (TBA) was monitored in real time by means of reflective interferometric spectroscopy (RIfS). The study has permitted to characterize in real time the path to immobilize TBA on the inner pore walls of NAA. Furthermore, this study provides an accurate label-free method to detect thrombin in real-time with high affinity and specificity.

In this work, a methodology that exploits the optical properties of the nanoporous anodic alumina gradient index filters (NAA-GIFs) has been developed and applied to evaluate in real time the release dynamics of a cargo molecule, acting as a model drug, filling the pores. NAA-GIFs with two photonic stopbands (PSBs) were prepared with one of its stop bands in the same absorption wavelength range of the cargo molecule, whereas the second stopband away from this absorption range. Numerical simulation and experiments confirm that the relative height of the high reflectance bands in the reflectance spectra of NAA-GIFs filled with the drug can be related to the relative amount of drug filling the pores. This property has been applied in a flow cell setup to measure in real-time the release dynamics of NAA-GIFs with the inner pore surface modified by layer-by-layer deposition of polyelectrolytes and loaded with the cargo molecule. The methodology developed in this work acts as a tool for the study of drug delivery from porous nanostructures. [ABSTRACT FROM AUTHOR]

The chemical modification, or functionalization, of the surfaces of nanomaterials is a key step to achieve biosensors with the best sensitivity and selectivity. The surface modification of biosensors usually comprises several modification steps that have to be optimized. Real-time monitoring of all the reactions taking place during such modification steps can be a highly helpful tool for optimization. In this work, we propose nanoporous anodic alumina (NAA) functionalized with the streptavidin-biotin complex as a platform towards label-free biosensors. Using reflective interferometric spectroscopy (RIfS), the streptavidin-biotin complex formation, using biotinylated thrombin as a molecule model, was monitored in real-time. The study compared the performance of different NAA pore sizes in order to achieve the highest response. Furthermore, the optimal streptavidin concentration that enabled the efficient detection of the biotinylated thrombin attachment was estimated. Finally, the ability of the NAA-RIfS system to quantify the concentration of biotinylated thrombin was evaluated. This study provides an optimized characterization method to monitor the chemical reactions that take place during the biotinylated molecules attachment within the NAA pores. [ABSTRACT FROM AUTHOR]