Álvaro Somoza

DNA‐Mediated Silver Nanoclusters: Synthesis, Properties and Applications

Fluorescent DNA-AgNCs have emerged as an alternative to standard emitters because of their unique properties: high fluorescent quantum yield, photostability, a broad pallet of colors (blue to near-IR), and the fact that their properties are easily modulated by the DNA sequence and environment. Applications as gene, ion, or small-molecule sensors have been reported.

Antibacterial Activity of DNA-Stabilized Silver Nanoclusters Tuned by Oligonucleotide Sequence

Silver nanoclusters (AgNCs) stabilized by DNA are promising materials with tunable fluorescent properties, which have been employed in a plethora of sensing systems. In this report, we explore their antimicrobial properties in Gram-positive and Gram-negative bacteria. After testing 9 oligonucleotides with different sequence and length, we found that the antibacterial activity depends on the sequence of the oligonucleotide employed. The sequences tested yielded fluorescent AgNCs, which can be grouped in blue, yellow, and red emitters. Interestingly, blue emitters yielded poor antibacterial activity, whereas yellow and red emitters afforded an activity similar to silver nitrate. Furthermore, structural studies using circular dichroism indicate the formation of complexes with different stability and structure, which might be one of the factors that modulate their activity. Finally, we prepared a trimeric structure containing the sequence that afforded the best antimicrobial activity, which inhibited the growth of Gram-positive and negative bacteria in the submicromolar range.

Multifunctionalization of magnetic nanoparticles for controlled drug release: a general approach.

In this study, a general approach for the multifunctionalization of magnetic nanoparticles (MNPs) with drugs (Doxorubicin and Gemcitabine) and targeting moieties (Nucant pseudopeptide) for controlled and selective release is described. The functionalization is achieved by the formation of disulfide bonds between MNPs and drugs derivatives synthesized in this work. Our strategy consists in the introduction of a pyridyldisulfide moiety to the drugs that react efficiently with sulfhydryl groups of pre-activated MNPs. This approach also allows the quantification of the covalently immobilized drug by measuring the amount of the 2-pyridinethione released during the process. The linkers developed here allow the release of drugs without any chemical modification. This process is triggered under highly reducing environment, such as that present inside the cells. Complete release of drugs is achieved within 5-8 h under intracellular conditions whereas negligible percentage of release is observed in extracellular conditions. We propose here a modular general approach for the functionalization of nanoparticles that can be used for different types of drugs and targeting agents.

Fluorescent DNA Stabilized Silver Nanoclusters as Biosensors

DNA stabilized fluorescent silver nanoclusters are promising materials, of which fluorescent properties can be exploited to develop sensors. Particularly, the presence of a DNA strand in the structure has promoted the development of gene sensors where one part of the sensor is able to recognize the target gene sequence. Moreover, since oligonucleotides can be designed to have binding properties (aptamers) a variety of sensors for proteins and cells have been developed using silver nanoclusters. In this review the applications of this material as sensors of different biomolecules are summarized.

Designed Modular Proteins as Scaffolds To Stabilize Fluorescent Nanoclusters.

Proteins have been used as templates to stabilize fluorescent metal nanoclusters thus obtaining stable fluorescent structures, and their fluorescent properties being modulated by the type of protein employed. Designed consensus tetratricopeptide repeat (CTPR) proteins are suited candidates as templates for the stabilization of metal nanoclusters due to their modular structural and functional properties. Here, we have studied the ability of CTPR proteins to stabilize fluorescent gold nanoclusters giving rise to designed functional hybrid nanostructures. First, we have investigated the influence of the number of CTPR units, as well as the presence of cysteine residues in the CTPR protein, on the fluorescent properties of the protein-stabilized gold nanoclusters. Synthetic protocols to retain the protein structure and function have been developed, since the structural and functional integrity of the protein template is critical for further applications. Finally, as a proof-of-concept, a CTPR module with specific binding capabilities has been used to stabilize gold nanoclusters with positive results. Remarkably, the protein-stabilized gold nanocluster obtained combines both the fluorescence properties of the nanoclusters and the functional properties of the protein. The fluorescence changes in nanoclusters fluorescence have been successfully used as a sensor to detect when the specific ligand was recognized by the CTPR module.

Glutathione-Triggered Drug Release from Nanostructures

The delivery of drugs can be improved with the use of different carriers, such as those based on nanoparticles. The nanostructures loaded with the therapeutic molecules should be able to reach the target cells and, what is more, release the drugs efficiently. Ideally, the drugs should be delivered only in the target cells, and not along their way to the cells. For these reasons several approaches have been developed to control the release of the drugs at the desired sites. In this review article we have summarized the reports that describe the use of glutathione to trigger the release of the therapeutic molecules from different nanostructures.

Single-point mutation detection in RNA extracts using gold nanoparticles modified with hydrophobic molecular beacon-like structures

Gold nanoparticles functionalized with oligonucleotides that bear a cholesterol group are used as gene sensors. The hydrophobic molecule is buried inside the nanostructure but when the complementary RNA sequence is present the structure unfolds exposing the cholesterol group to the water molecules. This rearrangement leads to the aggregation of the nanostructures.

Enhanced fluorescence of silver nanoclusters stabilized with branched oligonucleotides

DNA stabilized silver nanoclusters (AgNCs) are promising optical materials, whose fluorescence properties can be tuned by the selection of the DNA sequence employed. In this work we have used modified oligonucleotides in the preparation of AgNCs. The fluorescent intensity obtained was 60 times higher than that achieved with standard oligonucleotides.

Oligonucleotide Sensor Based on Selective Capture of Upconversion Nanoparticles Triggered by Target-Induced DNA Interstrand Ligand Reaction

We present a sensor that exploits the phenomenon of upconversion luminescence to detect the presence of specific sequences of small oligonucleotides such as miRNAs among others. The sensor is based on NaYF4:Yb,Er@SiO2 nanoparticles functionalized with ssDNA that contain azide groups on the 3′ ends. In the presence of a target sequence, interstrand ligation is possible via the click-reaction between one azide of the upconversion probe and a DBCO-ssDNA-biotin probe present in the solution. As a result of this specific and selective process, biotin is covalently attached to the surface of the upconversion nanoparticles. The presence of biotin on the surface of the nanoparticles allows their selective capture on a streptavidin-coated support, giving a luminescent signal proportional to the amount of target strands present in the test samples. With the aim of studying the analytical properties of the sensor, total RNA samples were extracted from healthy mosquitoes and were spiked-in with a specific target sequence at different concentrations. The result of these experiments revealed that the sensor was able to detect 10–17 moles per well (100 fM) of the target sequence in mixtures containing 100 ng of total RNA per well. A similar limit of detection was found for spiked human serum samples, demonstrating the suitability of the sensor for detecting specific sequences of small oligonucleotides under real conditions. In contrast, in the presence of noncomplementary sequences or sequences having mismatches, the luminescent signal was negligible or conspicuously reduced.

Gold-Coated Superparamagnetic Nanoparticles for Single Methyl Discrimination in DNA Aptamers

Au- and iron-based magnetic nanoparticles (NPs) are promising NPs for biomedical applications due to their unique properties. The combination of a gold coating over a magnetic core puts together the benefits from adding the magnetic properties to the robust chemistry provided by the thiol functionalization of gold. Here, the use of Au-coated magnetic NPs for molecular detection of a single methylation in DNA aptamer is described. Binding of α-thrombin to two aptamers conjugated to these NPs causes aggregation, a phenomenon that can be observed by UV, DLS and MRI. These techniques discriminate a single methylation in one of the aptamers, preventing aggregation due to the inability of α-thrombin to recognize it. A parallel study with gold and ferromagnetic NPs is detailed, concluding that the Au coating of FexOy NP does not affect their performance and that they are suitable as complex biosensors. These results prove the high detection potency of Au-coated SPIONs for biomedical applications especially for DNA repair detection.