Sensors based on upconverting nanoparticles for the detection of RNA/DNA oligolnucleotides

Tesis inédita de la Universidad Complutense de Madrid, Facultad de Farmacia, Departamento de Química en Ciencias Farmacéuticas, leída el 04-07-2019Depto. de Química en Ciencias FarmacéuticasFac. de FarmaciaTRUEunpu

Biohybrid sensor systems for the detection of metal ions in water

Die Wasserverschmutzung durch Seltenen Erden (REEs) und Schwermetallen verursacht viele Probleme für die Umwelt und die menschliche Gesundheit. Daher ist der Nachweis solcher Elemente von hoher Priorität. Derzeit verwendete Methoden haben einige Nachteile, wie hohe Messkosten, beschränke Selektivität, komplexe Handhabung oder der Bedarf von hochqualifiziertem Personal für die Probenanalyse. Die Kombination von biologischen Komponenten und Nanomaterialien zur Sensorentwicklung bietet eine Möglichkeit diese Nachteile ausgleichen. Mikroorganismen haben evolutionäre Strategien entwickelt, um sich vor toxischen Schwermetallen zu schützen, z.B durch Binden der Metallionen an ihrer Zelloberfläche mit speziellen Oberflächenproteinen (S-Layer). Diese bestehen aus einer Monolage identischer (Glyco-) Proteine, die sich selbst assemblieren und eine hochgeordnete kristalline Struktur unterschiedlicher Symmetrie bilden können. Studien haben die Bindung von Metallionen (einschließlich REEs) durch S-Layer-Proteine gezeigt. In dieser Dissertation wurden drei Nanomaterialien (Goldnanopartikel (AuNPs), planare Goldoberflächen und Nanodiamanten (NDs)) mit acht verschiedene S-Layer-Proteinen beschichtet. Ziel war die Entwicklung von Biohybrid-Sensor-Systemen für die Detektion von bis zu 12 Metallionen in Wasser. Ein kolorimetrisches Sensorsystem mit biofunktionalisierten AuNPs zur Detektion von REEs und Schwermetallen, einschließlich der aktuell vermehrt auftretenden Schadstoffe Lanthan und Gadolinium, wurde etabliert. Die Nachweisgrenzen lagen im Bereich vergleichbarer AuNPs-Systeme zum Nachweis von Schwermetallen, während die Slayer-AuNP-Biohybride ein breiteres Spektrum von Metallionen detektieren konnten. Das Screening aller acht S-Layer-AuNP-Biohybride mit 12 Metallsalzlösungen ergab charakteristische Wechselwirkungsmuster für jede der Kombinationen und ermöglichte den spezifischen Nachweis einer einzelnen Metallionenspezies in unbekannten Lösungen. Eine Kosten- und Ressourcenoptimierung ist über die Lagerung bis zu drei Monate und Wiederverwendbarkeit gegeben. Auf planaren Goldoberflächen ermöglichten die SPR-Spektroskopie die Messung der Adsorption von S-Layer-Proteinen, sowie die anschließende Detektion von CuSO4, NiCl2 und YCl3. Die Detektionslimits lagen dabei unter den kolorimetrischen Biohybridsystemen. Die SPR-Chips wurden erfolgreich regeneriert und für mehrere Funktionalisierungen mit S-Layer-Proteinen wiederverwendet. Das S-Layer-Protein SslA von S. ureae ATCC 13881 wurde erstmals an NDs adsorbiert. Die NDs/SslA-Biohybride wurden zur Detektion von CuCl2 und NiCl 2 verwendet, indem die Agglomeration und das Fluoreszenzquenching gemessen wurden. Es hat sich gezeigt, dass die vorgestellten Systeme viele der Nachteile ausgleichen, die mit derzeit verwendeten Systemen verbunden sind. Sie detektieren eine Vielzahl von Metallionen und minimieren so den Bedarf für mehrere Methoden. Die Nachweisgrenzen waren vergleichbar mit aktuellen kolorimetrischen und chemischen Kit-Systemen. Die S-layer-AuNPs und NDs/S-layer-Biohybride waren schnell und einfach zu handhaben, wodurch der Bedarf an hochqualifiziertem Messpersonal minimiert werden kann. Darüber hinaus führt die Verwendung von kostengünstigen Materialien wie NDs und die Wiederverwendbarkeit der Biohybride zu einem ressourceneffizienten und kostengünstigen Nachweissystem. Diese Dissertation hat das enorme Potenzial von S-Layer-Proteinen für den Nachweis von REEs und Schwermetallen in Wasser unter Verwendung verschiedener Nachweissysteme wie kolorimetrischer AuNPs-Assays, SPR-Spektroskopie und NDs gezeigt.The pollution of aqueous systems with rare earth elements (REEs) and heavy metals causes serious problems for environmental and human health. Therefore, the detection of such elements is of uttermost importance. Currently used methods have some disadvantages, such as high measurement costs, limited selectivity, complex sample handling, or the need for highly qualified personnel for sample analysis. The combination of biological components and nanomaterials for sensor development offers a way to offset these disadvantages. Microorganisms have developed strategies to protect themselves from heavy metal toxicity, e.g. by binding the metal ions on their cell surface with special Surface layer (S-layer) proteins. They consist of a monolayer of identical (glyco-) proteins, which can self-assemble and form a highly ordered crystalline structure of varying symmetry. Studies on the heavy metal binding of S-layer proteins have demonstrated their affinity for metal ions, including REE. The combination of nanomaterials with S-layer proteins enables the development of new sensors for these elements. Within this dissertation several nanomaterials in combination with S-layer proteins were investigated to obtain sensors for REEs and heavy metals. Eight different S-layer proteins were used to functionalize AuNPs, flat gold surfaces and nanodiamonds (NDs) for the detection of up to 12 metal ions in water. Colorimetric sensor systems with biofunctionalized AuNPs for the detection of REE and heavy metals, including the newly emerging pollutants lanthanum and gadolinium, were established. The detection limits of reference measurements and spiked tapwater samples were in the range of comparable AuNPs systems for the detection of heavy metals, while offering a broader range of metal ions to detect. The screening of all eight S-layer-AuNP biohybrids with 12 metal ions revealed specific interaction patterns for each of the combinations. The optimization cost and resource is achieved by storage up to three months and reusability of the S-layer-AuNP biohybrids. Surface plasmon resonance (SPR) spectroscopy enabled the measurement of S-layer proteins binding to flat gold surfaces, resulting in a stable protein layer used for the subsequent detection of CuSO4, NiCl2 and YCl3. The SPR chips were succesfully regenerated and reused for multiple functionalizations with S-layer proteins. The S-layer protein SslA from S. ureae ATCC 13881 was successfully adsorbed to the pristine NDs by physical conjugation. The NDs/SslA conjugates were used for the detection of CuCl2 and NiCl2, by measuring the agglomeration of the NDs and fluorescence quenching. The presented systems compensate many of the disadvantages associated with currently used techniques. They detect a broad variety of metal ions, minimizing the need for multiple methods. The detection limits were comparable to current colorimetric and chemical kit systems. The S-layer-AuNPs and NDs/S-layer biohybrids were quick and easy to handle, minimizing the need for highly qualitified personnel. In addition, the use of cost-effective materials such as NDs and the reusability of the biohybrids results in resource-efficient and cost-effective sensor systems. This project has shown the tremendous potential of S-layer proteins for the detection of REE and metal ions in water, by utilizing different detection systems like colorimetric AuNPs assays, SPR spectroscopy and NDs

Coupling colloidal chemistry with coordination chemistry: Design of hybrid nanomaterials by the assembly of plasmonic nanoparticles and functional coordination complexes

Nanotechnology involves the design, characterization, production and application of structures, devices and systems by the control of the shape and size at the nanometer scale involving different fields. In the last decade, nanotechnology development has boosted the interest in hybrid nanomaterials. These materials are a complimenting combination of two (or more) nanoparticles (NPs) with enhanced performance characteristics that offer exciting opportunities. It allows the possibility of integrating materials with different physical and chemical properties to widen the range of practical applications. In this context, Au NPs have recently attracted a lot of attention due to the great opportunities that Au offers at the nanoscale. In fact, their facile synthesis and functionalization can be exploited for constructing hybrid nanoparticles showing multi-functionality. In this manner, different Au hybrid nanostructures have been developed exhibiting diverse sizes, shapes and compositions displaying novel physicochemical properties, opening the door to potential new applications. On the other hand, Coordination Polymers (CPs) possess besides interesting electronic properties, potential advantages over conventional inorganic nanomaterials such as structural and chemical versatility, high specific area and biodegradability, among others. Therefore, the integration of both Au and CPs in a single heterostructure has emerged as an appealing topic. However, suitable chemical design appears as one of the key factors to improve their applicability. The work described in this thesis is motivated by the purpose of designing and studying novel hybrid nanostructures formed by combining Au NPs with different CPs: i) Prussian Blue and its Analogues (PB and PBA), ii) Spin-Crossover compounds (SCO) and iii) Metal-Organic Frameworks (MOF). Taking into account the numerous possible heterostructures, it will be discussed why these tailored hybrid NPs are the most appropriate for magneto-optical, electrochemical and electrical applications. In chapter 1, it is described the optical properties and the synthesis of Au NPs as well as the main research efforts that have been made to combine CPs incorporating Au functionalities within the overall hybrid nanomaterials. The main results of this thesis are divided into three parts depending on the potential applications: magneto-optics, electrochemistry and electrical conductivity. Chapter 2 deals with the preparation of hybrid systems formed by metallic NPs decorated by electrostatic attraction onto PBA NPs of different sizes and nature. In this approach, the capping agent of the plasmonic NP is modified, thus, allowing to select the plasmonic NP (isotropic or anisotropic) and, therefore, to tune the plasmon band position in a broad range of the visible spectrum. The heterostructure keeps its plasmonic and magnetic properties becoming a suitable hybrid material for magneto-optical applications. In chapter 3, different heterostructures composed of Au and PBA (of NiFe and CoFe) are synthesized and evaluated as electrocatalysts for the oxygen evolution reaction. The core@shell heterostructures are found to be the most appropriate to exploit the Au properties (conductivity and electronegativity). In this way, through a suitable chemical design it can be greatly enhanced the electrochemical activity and stability of the electroactive PBA. In chapter 4, a straightforward protocol is carried out to overgrow a thin SCO over different plasmonic NPs. Moreover, this synthetic route was extended to MOF. It is observed that thanks to the metallic core and the naked surface of the ultrathin SCO/MOF shell, these core@shell NPs are more conductive than the pristine SCO NPs when contacted to electrodes. In future work, further development will be done by taking advantage of the plasmon properties of the plasmonic core to get a light-induced spin transition (SCO) and to promote the adsorption/desorption of guest molecules (MOF) to obtain advanced sensing devices. This Ph.D. thesis is expected to represent a significant advancement in the development of novel heterostructures as a result of the incorporation of Au NPs to CPs

Façonnage de modes plasmons dans des colloïdes d'or

Cette thèse a pour objectif principal d'interpréter des mesures expérimentales des propriétés plasmoniques de nanoparticules d'or cristallines de différentes morphologies. Nous étudions tout d'abord, grâce à un film photosensible, la distribution du champ local autour de nano-prismes d'or. Cette cartographie indirecte révèle un effet de taille en irradiant à 457 nm et 514nm des objets de formes identiques mais de dimensions différentes, suggérant une réponse quasi-modale des plasmons supportés par ces structures. Cette réponse quasi-modale est mise en évidence dans le cas de prismes triangulaires de 500 nm de côté. Dans une seconde étape, nous montrons que la luminescence à deux photons (TPL) offre un autre moyen original de sonder la distribution du champ électromagnétique à l'intérieur de ces particules. En particulier, nous considérons le cas des nano-bâtonnets et des prismes d'or. Les cartes TPL montrent un fort confinement du champ à l'intérieur de ces objets. La localisation du signal peut être contrôlée en ajustant la polarisation incidente. Parallèlement, nous avons développé un nouvel outil de simulation pour reproduire et interpréter les cartes TPL. Elles confirment la spatialisation spécifique du champ proche et nous renseignent sur l'influence de la longueur d'onde et de la forme des prismes. Notre modèle suggère que la TPL donne aussi accès à la densité locale d'états plasmoniques dans la particule, et que le couplage entre deux nano-prismes peut constituer une nouvelle façon de concevoir des portes logiques modales plasmoniques. Enfin, nous avons étudié les propriétés optiques de chaines de sphères d'or d'une dizaine de nanomètres de diamètre. Les particules constituant ces chaines fusionnent sous l'effet du faisceau du microscope électronique en transmission ce qui conduit à un décalage dans l'infrarouge des modes de résonance plasmon. Nous avons caractérisé spatialement et spectralement ces modes plasmons par spectroscopie de pertes d'énergie (EELS). Grâce à la polyvalence du formalisme des fonctions de Green, et en nous appuyant sur une modélisation théorique montrant le lien entre signal de perte et densité locale d'états, nous avons simulé des cartes EELS de ces objets.The main objective of this thesis is to interpret experimental measurements of the plasmonic properties of gold colloidal particles. First, we study the optical near-field distribution around nano-platelets thanks to the photomigration technique. This indirect photo-chemical mapping uncovers a size effect by shining at 457 nm and 514 nm particles of different sizes, suggesting a quasi-modal response from the surface plasmons supported by these structures. This peculiar response is evidenced in the case of colloidal triangles of 500 nm side length. Then, we show that Two Photon Luminescence (TPL) offers another original way to probe the electromagnetic field distribution inside Au particles. We focus the TPL study on gold nano-rods and platelets. TPL maps show a strong confinement of the electromagnetic field inside these objects. Signal localization can be controlled by adjusting incident polarization. Simultaneously, we have developed a new tool based on the Green Dyadic Method (GDM) to reproduce and analyze TPL maps. They confirm specific near-field localization in the metal and provide a new insight into wavelength and particle shape influence. Our model suggests that TPL gives also access to the local density of plasmonic states in the particle and consequently that the coupling between two gold platelets could lead to a new design of a plasmonic modal logic gate. Finally, we have investigated the optical properties of gold nanoparticle chains. These nanoparticles fuse under the electron beam of the TEM. The fusion shifts plasmons resonances of the chains towards the infrared region. These plasmonic modes have been studied spatially and spectrally using Electron Energy Loss Spectroscopy (EELS). We have developed a theoretical model showing the link between the energy loss of a swift electron and the local density of states. Using the GDM, we have computed EELS maps of these nanoparticle chains