This dissertation is about fabrication and functionalization of metal nanoparticles for use in plasmonic and nonlinear optical (NLO) applications. In the first two chapters, I describe a series of experiments, where I combined silver nanoparticles fabricated by nanosphere lithography with ionic self-assembled multilayer (ISAM) films, tuning the geometry of the particles to make their plasmonic resonances overlap with the frequency of optical excitation. The designed hybrid metallic/organic nanostructures exhibited large enhancements of the efficiency of second harmonic generation (SHG) compared to conventional ISAM films, causing a modified film with just 3 bilayers to be optically equivalent to a conventional 700-1000 bilayer film. SHG responses from Ag nanoparticle-decorated hybrid-covalent ISAM (HCISAM) films were investigated as the next logical step towards high-Ï ²⁺ ISAM films. I found that the plasmonic enhancement primarily stems from interface SHG. Interface effects were characterized by direct comparison of SHG signals from PAH/PCBS ISAM films and PAH/PB HCISAM films. Though interface &chi²⁺ is substantially smaller in PAH/PCBS than in PAH/PB, plasmonically enhanced PAH/PCBS films exhibit stronger NLO response. I propose that the structure of PAH/PB film makes its interface more susceptible to disruptions in the nanoparticle deposition process, which explains our observations. During the fabrication of monolayer crystals for nanosphere lithography, I developed a variation of the technique of convective self-assembly, where the drying meniscus is restricted by a straight-edge located approximately 100 μM above the substrate adjacent to the drying zone. This technique can yield colloidal crystals at roughly twice the growth rate compared to the standard technique. I attribute this to different evaporation rates in the thin wet films in the two cases. I also found that the crystal growth rate depends strongly on the ambient relative humidity. Finally, dithiocarbamate (DTC)-grafted polymers were synthesized and employed to functionalize surfaces of Au nanopartciles. PAH-DTC shows greater stability in different environments than PEI-DTC. I also investigated the stability of PAH-DTC coated particles in suspensions with UV-Vis spectroscopy and autotitration. The covalently bonded PAH-DTC enhances the colloidal stability of the Au nanoparticles and enables subsequent ISAM film deposition onto the particles. / Ph. D.

The environmental fate and transport of engineered nanomaterials have been broadly investigated and evaluated in many published studies. Silver nanoparticles (AgNPs) represent one of the most widely manufactured nanomaterials. They are currently being incorporated into a wide range of consumer products due to their purported antimicrobial properties. However, either the AgNPs themselves or dissolved Ag+ ions has a significant potential for the environmental release. The safety issues for nanoparticles are continuously being tested because of their potential danger to the environment and human health. Studies have explored the toxicity of AgNPs to a variety of organisms and have shown such toxicity is primarily driven by Ag+ ion release. Dissolution of nanoparticles is an important process that alters their properties and is a critical step in determining their safety. Therefore, studying nanoparticles' dissolution can help in the current move towards safer design and application of nanoparticles. This research endeavor sought to acquire comprehensive kinetic data of AgNP dissolution to aid in the development of quantitative risk assessments of AgNP fate. To evaluate the dissolution process in the absence of nanoparticle aggregation, AgNP arrays were produced on glass substrates using nanosphere lithography (NSL). Changes in the size and shape of the prepared AgNP arrays were monitored during the dissolution process by atomic force microscopy (AFM). The dissolution of AgNP is affected by both internal and external factors. First, surface coating effects were investigated by using three different coating agents (BSA, PEG1000, and PEG5000). Capping agent effects nanoparticle transformation rate by blocking reactants from the nanoparticle surface. Coatings prevented dissolution to different extents due to the various way they were attached to the AgNP surface. Evidence for the existence of bonds between the coating agents and the AgNPs was obtained by surface enhanced Raman spectroscopy. Moreover, to study the size effects on AgNP dissolution, small, medium, and large sized AgNPs were used. The surrounding medium and temperature were the two variables that were included in the size effects study. Relationships were established between medium concentration and dissolution rate for three different sized AgNP samples. By using the Arrhenius equation to plot the reaction constant vs. reaction temperature, the activation energy of AgNPs of different sizes were obtained and compared. / Doctor of Philosophy / Nanomaterials, defined as materials with at least one characteristic dimension less than 100 nm, often have useful attributes that are distinct from the bulk material. The novel physical, chemical, and biological properties enable the promising applications in various manufacturing industry. Silver nanoparticles (AgNPs) represent one of the most widely manufactured nanomaterials and has been used as the antimicrobial agent in a wide range of consumer products. However, either the AgNPs themselves or dissolved Ag+ ions has a significant potential for the environmental release. The environmental fate and transport of AgNPs drawn considerable attentions because of the potential danger to environment and human health. Dissolution of nanoparticles is an important process that alters their properties and is a critical step in determining their safety. Ag+ ions migrate from the nanoparticle surface to the bulk solution when an AgNP dissolves. Studying nanoparticles' dissolution can help in the current move towards safer design and application of nanoparticles. This research aimed to acquire comprehensive kinetic data of AgNP dissolution to aid in the development of quantitative risk assessments of AgNP fate. AgNP arrays were produced on glass substrates using nanosphere lithography (NSL) and changes in the size and shape during the dissolution process were monitored by atomic force microscopy (AFM). First, surface coating effects were investigated by using three different coating agents. Coatings prevented dissolution to different extents due to the various way they were attached to the AgNP surface. Moreover, small, medium, and large sized AgNPs were used to study the size effects on AgNP dissolution. The surrounding medium concentration and temperature were the two variables that were included in the size effects study.

Whole-cell sensors represent an emerging branch in biosensor development since they obviate the need for enzyme/antibody purification and provide the unique opportunity to assess global parameters such as genotoxicity and bioavailability. Yeast species such as Saccharomyces cerevisiae are ideal hosts for whole-cell sensor applications. However, current approaches almost exclusively rely on analyte-induced expression of fluorescent proteins or luciferases that imply issues with light scattering and/or require the supply of additional substrates. In this study, the yeast α-factor mating pheromone, a peptide pheromone involved in cell-cell communication in Saccharomyces cerevisiae, was utilized to create the whole-cell sensor read-out signal, in particular by employing engineered sensor cells that couple the response to a user-defined environmental signal to α-factor secretion. Two novel immunoassays - relying on hydrophobin-based surface engineering - were developed to quantify the α-factor. Hydrophobins are amphiphilic fungal proteins that self-assemble into robust monolayers at hydrophobic surfaces. Two recombinant hydrophobins, either lacking (EAS) or exposing the α-factor pheromone (EAS-α) upon self-assembly, were used to functionalize polystyrene supports. In a first approach (competitive immunoassay), pheromone-specific antibodies initially bound to the functionalized surface (due to the α-factor exposed by the hydrophobin layer) were competitively detached by soluble α-factor. In a second approach, the antibodies were first premixed with pheromone-containing samples and subsequently applied to functionalized surfaces, allowing for the attachment of antibodies that still carried available binding sites (inverse immunoassay). Both immunoassays enabled quantitative assessment of the yeast pheromone in a unique but partially overlapping dynamic range and allowed for facile tuning of the assay sensitivity by adjustment of the EAS-α content of the hydrophobin layer. With a limit of detection of 0.1 nM α-factor, the inverse immunoassay proved to be the most sensitive pheromone quantification assay currently available. Due to the high stability of hydrophobin monolayers, functionalized surfaces could be reused for multiple consecutive measurements. Favorably, both immunoassays proved to be largely robust against the changes in the sample matrix composition, allowing for pheromone quantification in complex sample matrices such as yeast culture supernatants. Hence, these immunoassays could also be applied to study the pheromone secretion of wild-type and engineered Saccharomyces cerevisiae strains. Additionally, a proof-of-concept whole-cell sensor for thiamine was developed by combining the hydrophobin-based immunoassays with engineered sensor cells of Schizosaccharomyces pombe modulating the secretion of the α-factor pheromone in response to thiamine. Since this read-out strategy encompasses intrinsic signal amplification and enables flexible choice of the transducer element, it could contribute to the development of miniaturized, portable whole-cell sensors for on-site application.

Iron oxide magnetic nanoparticle synthesis and their surface functionalization hold a crucial position in the design and fabrication of functional materials for a variety of biomedical applications. Non-uniform nanoparticles with poor crystallinity, prepared by conventional methods, have only limited value in biological areas. Large scale synthesis methods that are able to produce high quality, mono-dispersed iron oxide nanoparticles using low cost and environment friendly chemicals are highly desirable. Following synthesis, appropriate surface functionalization is necessary to direct the dispersibility of nanoparticles in aqueous solution in order to provide them with acceptable colloidal stability against the ion strength and many biomolecules that nanoparticles may encounter under physiological conditions. Poorly stabilized nanoparticles that easily aggregate and form large size agglomerates would be quickly cleared by the liver and other organs and are not suitable for clinical purposes. Additionally, many interesting functionalities such as fluorescence, targeting and anti-cancer properties can be immobilized onto the surface of iron oxide magnetic nanoparticles during the surface functionalization process so as to build multifunctional platforms for disease diagnosis and treatment. Polyol method can be an effective way to prepare magnetite nanoparticles that are suitable for biological applications. In a polyol system, selected surface functionalities were introduced to the nanoparticle surface via a hot injection technique. The morphology, uniformity, crystallinity and magnetic properties were examined to understand the effect of different ligand molecules on the final product. Their surface chemistry, colloidal properties and surface reactivity were also studied to evaluate their practicability in different applications. A high efficiency in-situ method for the preparation of magnetite nanoparticles attached to silica nanospheres was also developed in a polyol system. This approach eliminates several time-consuming processing steps that are in the conventional fabrication route and directly produces water-stable magnetite-silica hybrid materials with surface availability for subsequent modifications. In addition to polyalcohol, the potential of polyamine in the preparation of water-soluble magnetite nanoparticles with amine surface functionalities was also evaluated. And it is suggested that polyamine acts as solvent, stabilizing agent and reducing agent simultaneously during the synthesis. The characterization of polyamine coated nanoparticles, their surface functionalization, and subsequent application for bioseparation and drug delivery were reported.

Dans ce travail de thèse, nous avons développé un nouveau procédé de production et de purification en temps réel des domaines 9 et 10 de la fibronectine humaine chez E.coli. Cette stratégie combine trois partenaires de fusion en tandem : un double tag d’affinité (10xHis et SBP) et un tag de coloration (le domaine de fixation de l’hème du cytochrome b5) en présence d’un site Tev. Ce système a montré sa performance dans le traçage visuel des étapes d’expression et de purification et dans la quantification de la CMAT-FNIII9-10. La présence du double tag d’affinité permet une étape de purification simple et offre un degré de pureté atteignant les 98%. Par ailleurs, le cytochrome b5 a montré son intérêt dans le suivi visuel et quantitatif de l’adsorption de la CMAT-FNIII9-10 à la surface d’un support plastique. Ensuite, l’activité du fragment recombinant a été validée avec succès. Dans cette étude, nous avons construit une matrice adhésive en combinant les propriétés du polymère PCL avec celles de la CMAT-FNIII9-10. L’immobilisation de celle-ci s’est opérée d’une manière orientée en adsorbant la protéine de fusion à la surface des PCL par l’intermédiaire de la streptavidine. Cette approche a conduit à l’élaboration d’un matériau biofonctionnalisé en optimisant l’exposition des sites d’attachement cellulaire à la surface des PCL par une immobilisation orientée. La réponse cellulaire des CMS humaines a été validée efficacement sur cette matrice, en absence de sérum et en présence de BSA. Les résultats de cette expérience montrent que cette stratégie a contribué à améliorer l’exposition des sites « RGD » et « PHSRN » favorisant l’interaction avec les récepteurs cellulaires. / In this thesis, we have developed a novel method for producing and purifying the 9th and 10thdomains of type III human fibronectin in E.coli. This strategy combines three fusion partnersin tandem: a dual affinity tag (10xHis and SBP) and a coloring tag (the binding domain of cytochrome b5 heme) in the presence of a Tev cleaving site.This system has demonstrated its performance in the visual tracking of the expression,purification and quantification steps of CMAT-FNIII9-10. The presence of the dual affinity tag allows a simple purification step and offers a degree of purity up to 98%. Moreover, the cytochrome b5 showed its interest in the visual and quantitative monitoring of the CMATFNIII9-10 adsorption onto the surface of a plastic support. Then the biological activity of there combinant fragment was successfully validated. In this study, we constructed an adhesive matrix by combining the properties of PCL with those of CMAT-FNIII9-10. Immobilization of the recombinant fragments is carried out by an oriented adsorption of the fusion protein onto the PCL film through a streptavidin layer. This approach has led to the development of a bio-functionalized material by optimizing the exposure of cell attachment sites on the surface of the PCL by an oriented immobilization. The cellular response of human MSCs was effectively validated on this matrix, in the absence of serum and in the presence of BSA. The results of this experiment show that this strategy has helped to improve the exposure sites "RGD" and "PHSRN" promoting interaction with cellular receptors.

Le marché du diagnostic connait un essor considérable depuis l’avènement de labiologie moléculaire. Plus précis et souvent plus rapide, le diagnostic moléculaire in vitro(DIV) est de plus en plus utilisé dans les laboratoires d’analyses médicales. L’ensemble destests dédiés au marché du DIV répond à des contraintes socio-économiques très précisescomme : la fiabilité du résultat, le délai de réponse court, le faible coût et la facilitéd’utilisation. Les indicateurs socio-économiques montrent que la technologie des biopuces estun potentiel bon candidat pour répondre aux attentes du marché. En effet, cet outil permetl’analyse simultanée de plusieurs dizaines voire centaines de séquences nucléiques et doncl’identification d’autant d’organismes en une seule analyse. Cette technologie s’inscrit encomplément de la PCR en apportant l’avantage de l’analyse multiplexée à moyen débit. Deplus, elle permet de donner une réponse globale de la multiplicité des espèces présentes dansl’échantillon sans avoir besoin de passer par une étape de culture. Néanmoins, cettetechnologie n’est pas optimisée pour le marché du DIV. En effet, son usage est complexe, peurobuste et trop cher pour concurrencer les méthodes actuelles (microbiologie pasteurienne,PCR, Elisa, etc..). Dans le but de réduire le coût de fabrication des biopuces à ADN, il estdonc nécessaire de développer des méthodes alternatives. Dans un premier temps, l’objectif de cette thèse Cifre a été de mettre au point unprototype nouveau de dépôt de biomolécules basé sur la lithographie douce, permettant dedéposer les oligonucléotides sondes de façon multiplexée et selon des motifs micrométriques.Cette nouvelle technologie a été évaluée par rapport aux technologies de références. Puis,nous avons développé un procédé innovant de double fonctionnalisation de surface. Ceprocédé simple et rapide a pour avantages de fonctionnaliser la biopuce avec la chimie desurface et les sondes en une seule étape et d’augmenter les signaux d’hybridation. La secondepartie de la thèse a été de coupler cette nouvelle technologie à la détection des événementsd’hybridation sans marquage en utilisant la diffraction de la lumière. La principale différenceavec la méthode de détection par fluorescence repose sur l’adressage des sondes. En effet, ledépôt doit être réalisé sous forme de réseaux de lignes nanométriques diffractants de façon àce que l'interaction entre les molécules déposées et les cibles qui interagissent soit trèssensible. Cette seconde phase du projet a été très ambitieuse et innovante. La faisabilité decette méthode de détection, démontrée par des simulations théoriques, a fait l’objet d’untravail d’optimisation très important et les résultats obtenus montrent que cette technologiesans marquage est possible. / The diagnosis market increased since the advent of molecular biology. More precise and often faster, the in vitro molecular diagnosis (DIV) is more and more used in medical analyses laboratories. DNA chips technology seems to be a good candidate to answer the market expectations. Indeed, this tool allows making several hundreds of analyses simultaneously. Furthermore, it allows giving a global answer of all the present species in the sample without the need of a culture step. Nevertheless, this technology is not optimized for the market of the DIV. Indeed, its use is complex and too expensive in comparison with the current methods (Pasteur microbiology, PCR, Elisa, etc.). So it is necessary to develop an alternative method to reduce the manufacturing cost and simplify the use of DNA chips. First, the goal of this industrial PhD Cifre supported by the Dendris Company was to complete a new prototype of biomolecules deposition based on soft lithography, allowing multiplexing the deposition of oligonucleotides probes along micro and nanometric patterns.This new technology was compared with the reference technologies. Then, we developed an innovative process of surface co-functionalization. This simple and fast process permits to functionalize the DNA chips with both surface chemistry and probes in a single step and to increase the hybridization signals. The second part of this PhD thesis was to couple this new technology with label-free detection using light diffraction. The main difference with fluorescence-based detection was about probes patterning. Indeed, we needed to generate molecular gratings of nanometric lines to diffract efficiently light from a laser beam. We showed that the absolute diffraction intensity increase with the gratings thickness, which is directly correlated with, probes and targets interactions. The second phase of the project was very ambitious and innovative because we demonstrated the feasibility of this label-free detection. And now we can think that this technology will appear as an alternative method for the diagnosis

The aim of biomaterials design is to create an artificial environment that mimics the in vivo extracellular matrix for optimized cell interactions. A precise synergy between the scaffolding material, bioactivity, and cell type must be maintained in an effective biomaterial. In this work, we present a technique of nanofabrication that creates chemically nanopatterned bioactive silicon surfaces for cell studies. Using nanoimprint lithography, RGD and mimetic BMP-2 peptides were covalently grafted onto silicon as nanodots of various dimensions, resulting in a nanodistribution of bioactivity. To study the effects of spatially distributed bioactivity on cell behavior, mesenchymal stem cells (MSCs) were cultured on these chemically modified surfaces, and their adhesion and differentiation were studied. MSCs are used in regenerative medicine due to their multipotent properties, and well-controlled biomaterial surface chemistries can be used to influence their fate. We observe that peptide nanodots induce differences in MSC behavior in terms of cytoskeletal organization, actin stress fiber arrangement, focal adhesion (FA) maturation, and MSC commitment in comparison with homogeneous control surfaces. In particular, FA area, distribution, and conformation were highly affected by the presence of peptide nanopatterns. Additionally, RGD and mimetic BMP-2 peptides influenced cellular behavior through different mechanisms that resulted in changes in cell spreading and FA maturation. These findings have remarkable implications that contribute to the understanding of cell-extracellular matrix interactions for clinical biomaterials applications.

Nano-objects have unique properties due to their sizes, shapes and structure. When electronic properties of such nano-objects are used to build devices, the control of interfaces at atomic level is required. In this thesis, systems were built that can not only electrically characterize nano-objects, but also allow to analyze a large number of individual nano-objects statistically at the example of graphene and nanoparticle-molecule-nanoelectrode junctions. An in-situ electrical characterization system was developed for the analysis of free standing graphene sheets containing defects created by an acid treatment. The electrical characterization of several hundred sheets revealed that the resistance in acid treated graphene sheets decreased by 50 times as compared to pristine graphene and is explained by the presence of di-vacancy defects. However, the mechanism of defect insertion into graphene is different when graphene is bombarded with a focused ion beam and in this case, the resistance of graphene increases upon defect insertion. The defect insertion becomes even stronger at liquid N2 temperature. A molecular electronics platform with excellent junction properties was fabricated where nanoparticle-molecule chains bridge 15-30nm nanoelectrodes. This approach enabled a systematic evaluation of junctions that were assembled by functionalizing electrode surfaces with alkanethiols and biphenyldithiol. The variations in the molecular device resistance were several orders of magnitude and explained by variations in attachment geometries of molecules.  The spread of resistance values of different devices was drastically reduced by using a new functionalization technique that relies on coating of gold nanoparticles with trityl protected alkanedithiols, where the trityl group was removed after trapping of nanoparticles in the electrode gap. This establishment of a reproducible molecular electronics platform enabled the observation of vibrations of a few molecules by inelastic tunneling spectroscopy. Thus this system can be used extensively to characterize molecules as well as build devices based on molecules and nanoparticles.

La détection et la surveillance des gaz est un enjeu important tant pour la sécurité industrielle que pour la protection de l’environnement et des personnes. Le dihydrogène, prend une place de plus en plus importante en tant que combustible et vecteur énergétique mais il est extrêmement inflammable et explosif dans un large domaine de 4 à 75 % dans l’air. De même, l’ammoniac est très utilisé dans l’industrie comme gaz réfrigérant ou comme élément de base pour la production chimiques d’autres composés. Ce gaz présente des risques sur l’environnement et sur les êtres vivants et peut former des mélanges explosifs avec l’air dans les limites de 15 à 28 % en volume. Les capteurs de gaz permettant d’indiquer la présence et/ou la quantification de ces gaz prennent alors toute leur importance. Dans la continuité de nos nombreux travaux sur les capteurs résistifs à base d’assemblages discontinus de nano-objets, l’objet de ce travail de thèse a été de préparer des capteurs résistifs pour la détection de H2 et NH3. Ces capteurs sont à base d’assemblages 2D de nanoparticules de compositions complexes. Trois types de nanoparticules cœur-coquille ont été synthétisés : Au@ZnO, Au@SnO2 et Au@Ag. Différentes techniques physico-chimiques (UV-Visible/TEM / DRX etc) ont permis de caractériser les particules obtenues. L’étape suivante a consisté à les assembler en monocouches compactes. Les films ont été obtenus par la méthode d’assemblage de Langmuir-Blodgett. Après transfert à la surface d’un substrat en verre supportant des électrodes inter digitées, les performances de détection des capteurs résistifs fabriqués ont été alors évaluées. Les capteurs à base de Au@ZnO et Au@SnO2 ont été testés sous H2, tandis que les capteurs à base de Au@Ag l’ont été sous NH3. Les capteurs fabriqués ont montré des performances attractives de détection de H2 et NH3 dans des gammes de concentration étendues. Une autre contribution importante de ce travail concerne la compréhension des mécanismes de détection. Diverses techniques analytiques, tels que la TPD (Température désorption Programmed) et la TPR (Température de réduction programmée) ont été utilisés pour permettre la discussion des les mécanismes impliqués. / Gas sensing and monitoring are important issues for both industrial safety and protection of the environment and human beings. Dihydrogen, is increasingly used as fuel and energy carrier but it is extremely flammable and explosive in a wide range between 4 and 75% in air.Similarly, ammonia is widely used in industry as a cooling gas or as a reagent for the chemical production of other compounds.This gas presents risks to the environment and to living beings and can form explosive mixtures with air within 15 to 28% by volume.Gas sensors, indicating the presence and /or quantification of these gases, are very important.In continuation of our work on resistive sensors based on discontinuous assembly of nano-objects, the aim of this thesis was to prepare resistive sensors for the detection of H2 and NH3.These sensors are based on 2D assemblies of complex compositions of nanoparticles. Three types of core-shell nanoparticles were synthesized: Au@ZnO, Au@SnO2 and Au@Ag. Different physicochemical techniques (UV-Visible / TEM / DRX etc.) were used to characterize the particles. The next step was to assemble them in compact monolayers. The films were obtained by Langmuir-Blodgett assembling technique. Then, they were transferred to the surface of a glass slide supporting interdigitated electrodes. Sensing performances of the as-fabricated resistive sensor were evaluated.Sensors based on Au@ZnO and Au@SnO2 nanoparticles were tested towards H2, while Au@Ag based sensors were tested under NH3.The sensors showed attractive performances in H2 and NH3 detection within wide concentration ranges. Another important contribution of this work is the understanding of detection mechanisms. Various analytical techniques such as TPD (Temperature Programmed Desorption) and TPR (temperature programmed reduction) were used for the discussion of the mechanisms involved.

La thèse commence le 1er octobre 2012. IL s'agit pour cette première année de thèse de synthétiser différents complexes de coordination a base du Ruthénium, Fer, Manganèse et de Cobalt à ligands polypyridinique. Ces complexes seront ensuite assemblés par lien covalent. Nous proposons d'utiliser entre autre les techniques d'electrochimie pour induire ces couplages. On cherchera aussi a immobiliser les assemblages sur surface. La principale difficulté attendue dans cette etape du travail concerne la fonctionalisation dissymétrique des ligands pour l'obtention de triade. / This thesis deals with the synthesis and characterization of inorganic triads for photoinduced charge separation. A range of photosensitizers will be studied, and the effect of the donor and acceptor will be assesed as well. These systems will be studied in solution and immobalized on surfaces. Cette thèse traite de la synthèse et la caractérisation des triades inorganiques pour la séparation de charge photoinduite. Une gamme de photosensibilisateurs seront étudiées, et l'effet du donneur et accepteur sera ainsi étudié. Ces systèmes seront étudiés en solution et immobalizes sur les surfaces.