Selective and Sensitive Probe Based in Oligonucleotide-Capped Nanoporous Alumina for the Rapid Screening of Infection Produced by Candida albicans.

A robust, sensitive, and time-competitive system to detect Candida albicans in less than 30 min in clinical samples based in capped nanoporous anodic alumina (NAA) is developed. In the proposed design, NAA pores are loaded with rhodamine B and then blocked with an oligonucleotide that is able to recognize C. albicans DNA. The capped material shows negligible cargo release, whereas dye delivery is selectively accomplished when genomic DNA from C. albicans is present. This procedure has been successfully applied to detect C. albicans in clinical samples from patients infected with this yeast. When compared with classical C. albicans detection methods, the proposed probe has a short assay time, high sensitivity and selectivity, demonstrating the high potential of this simple design for the diagnosis of infection produced by C. albicans.

Nanoporous Anodic Alumina Surface Modification by Electrostatic, Covalent, and Immune Complexation Binding Investigated by Capillary Filling.

The fluid imbibition-coupled laser interferometry (FICLI) technique has been applied to detect and quantify surface changes and pore dimension variations in nanoporous anodic alumina (NAA) structures. FICLI is a noninvasive optical technique that permits the determination of the NAA average pore radius with high accuracy. In this work, the technique is applied after each step of different surface modification paths of the NAA pores: (i) electrostatic immobilization of bovine serum albumin (BSA), (ii) covalent attachment of streptavidin via (3-aminipropyl)-triethoxysilane and glutaraldehyde grafting, and (iii) immune complexation. Results show that BSA attachment can be detected as a reduction in estimated radius from FICLI with high accuracy and reproducibility. In the case of the covalent attachment of streptavidin, FICLI is able to recognize a multilayer formation of the silane and the protein. For immune complexation, the technique is able to detect different antibody-antigen bindings and distinguish different dynamics among different immune species.

Molecular gated nanoporous anodic alumina for the detection of cocaine.

We present herein the use of nanoporous anodic alumina (NAA) as a suitable support to implement "molecular gates" for sensing applications. In our design, a NAA support is loaded with a fluorescent reporter (rhodamine B) and functionalized with a short single-stranded DNA. Then pores are blocked by the subsequent hybridisation of a specific cocaine aptamer. The response of the gated material was studied in aqueous solution. In a typical experiment, the support was immersed in hybridisation buffer solution in the absence or presence of cocaine. At certain times, the release of rhodamine B from pore voids was measured by fluorescence spectroscopy. The capped NAA support showed poor cargo delivery, but presence of cocaine in the solution selectively induced rhodamine B release. By this simple procedure a limit of detection as low as 5 × 10-7 M was calculated for cocaine. The gated NAA was successfully applied to detect cocaine in saliva samples and the possible re-use of the nanostructures was assessed. Based on these results, we believe that NAA could be a suitable support to prepare optical gated probes with a synergic combination of the favourable features of selected gated sensing systems and NAA.

In Vitro Biocompatibility of Surface-Modified Porous Alumina Particles for HepG2 Tumor Cells: Toward Early Diagnosis and Targeted Treatment.

Porous alumina photoluminescence-inherent particles are produced and proposed for the development of biomarkers detectors and localized treatment of HepG2 cells. Nanoporous alumina particles (NPAPs) are amorphous, consist of hexagonally ordered nanometric pores in an alumina matrix, have high chemical stability in physiological pH, and exhibit a high inherent photoluminescence in the visible spectrum independently of their size, selectable from nanometers to tens of micrometers. The surface of NPAPs is chemically modified using two different functionalization methods, a multistep method with (3-aminopropyl)triethoxysilane (APTES) and glutaraldehyde (GLTA) and a novel simplified-step method with silane-PEG-NHS. Fourier Transform infrared spectroscopy analysis confirmed the proper surface modification of the particles for both functionalization methods. HepG2 cells were cultured during different times with growing concentrations of particles. The analysis of cytotoxicity and cell viability of HepG2 cells confirmed the good biocompatibility of NPAPs in all culture conditions. The results prove the suitability of NPAPs for developing new label-free biomarker detectors and advantageous carriers for localized drug delivery.

Protein attachment to nanoporous anodic alumina for biotechnological applications: influence of pore size, protein size and functionalization path.

Nanoporous anodic alumina (NAA) is a material with great interest in nanotechnology and with promising applications to biotechnology. Obtaining specific and regularly functionalized NAA surfaces is essential to obtain meaningful results and applications. Silane-PEG-NHS (triethoxysilane-polyethylene-glycol-N-hydroxysuccinimide) is a covalent linker commonly used for single-molecule studies. We investigate the functionalization of NAA with silane-PEG-NHS and compared with two common, but not single-molecule, grafting agents, APTMS (3-aminopropylotrimethoxysilane) as an electrostatic linker, and APTMS-GTA (3-aminopropylotrimethoxysilane-glutaraldehyde) as covalent. Another outcome of this study is to show how two proteins (collagen and bovine serum albumin, BSA) with different properties differentially arrange for different functionalizations and NAA pore sizes. FTIR is used to demonstrate the surface modification steps and fluorescence confocal microscopy reveals that silane-PEG-NHS results in a more homogeneous protein distribution in comparison to the other linkers. Reflection interference Fourier transform spectroscopy confirms the confocal fluorescence microscopy results and permits to estimate the amounts of linker and linked proteins within the pores. These results permit to obtain uniformly chemical modified NAA supports with a great value in biosensing, drug delivery and cell biology.