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With the aim of developing a deeper understanding of particle behaviour within nano-hybrid materials, this thesis investigates the particle-particle and particle-polymer interactions that influence and control cellulose nanocrystal dispersion in aqueous and non-aqueous environments. / Cellulose nanocrystals (CNCs) are rigid rod-shaped nanoparticles derived from bio-based resources and are considered an emerging nanomaterial based on their commercial availability and favourable properties. CNCs have great potential as reinforcing agents in hybrid materials and composite applications if they are well-dispersed. Whereas colloidal stability is effectively described by established theories, dispersing nanoparticles from an aggregated state, and their interaction with polymers can be difficult to predict and control. Herein, the particle-particle and particle-polymer interactions that govern CNC dispersibility in aqueous and non-aqueous environments are examined. The surface chemistry, morphology and colloidal/thermal stability of CNCs from North American industrial producers were extensively characterized such that particle interactions could be reproducibly measured from a known starting material. Industrially produced CNCs compared well to those produced at the bench-scale, implying that laboratory results should be translatable to the development of new CNC-based products. To examine particle-particle interactions within dry CNC aggregates, a surface plasmon resonance-based platform was developed to monitor CNC film swelling in a range of solvents and salt solutions. Water was observed to interrupt particle-particle hydrogen bonding most effectively, however film stability, and ultimately particle aggregation, was maintained by strong van der Waals interactions. Moreover, particle spacing and overall film thickness was found to be independent of the CNC surface chemistries and surface charge densities examined, yet the rate of film swelling scaled with the ionic strength of the surrounding media. Polyethylene glycol (PEG) was used as a model, non-ionic, water-soluble polymer to investigate polymer adsorption to CNC surfaces in water. PEG did not adsorb to CNCs despite the abundance of hydroxyl groups, which is in direct contrast to silica particles that are well known to hydrogen bond with PEG. Combining the knowledge of both particle-particle and particle-polymer interactions, PEG nanocomposites reinforced with CNCs and silica were compared and particle dispersibility was related to composite performance. Although PEG does not adsorb to CNCs in aqueous environments, polymer adsorption does occur in dry polymer nanocomposites leading to good dispersibility and improved mechanical properties. Overall, the work presented here yields new insight into the forces that govern CNC dispersion and provides a foundation from which a variety of new CNC-based products can be developed. / Thesis / Doctor of Philosophy (PhD) / Using particles derived from renewable resources to reinforce plastics and other materials has the potential to make products lighter, stronger and more environmentally friendly. However, to make these products we need to understand how to control and distribute particles uniformly throughout hybrid/composite materials. This work uses particles extracted from trees and cotton, known as cellulose nanocrystals, to reveal which factors govern particle dispersion in reinforced composite materials. To do so, first the properties and performance of commercially available cellulose nanocrystals were extensively analyzed and compared to form the basis from which interactions can be understood. Next, particle films were measured in water, organic solvents and salt solutions to better understand how aggregated cellulose nanocrystals can be separated within composite materials. The interactions between water-soluble polymers and cellulose nanocrystals were then investigated to reveal how polymer adsorption impacts particle dispersibility. Finally reinforced polymer composites were prepared with uniformly distributed cellulose nanocrystals and the crystallization and mechanical properties were investigated. By developing a deeper understanding of the factors that control cellulose nanocrystal dispersion we can learn how to make a variety of new and improved environmentally conscious products.

Micro and nanofabrication technologies have been used extensively in many biomedical and biological applications. Integration of MEMS technology and biology (BioMEMS) enables precise control of the cellular microenvironments and offers high throughput systems. The focus of this research was to develop three dimensional (3-D) isotropic microstructures for comprehensive analysis on cell-substrate interactions. The aim was to investigate whether the normal and cancerous cells differentially respond to their underlying substrate and whether the differential response of the cells leads to a novel label-free technique to distinguish between normal and cancerous cells. Three different generations of 3-D isotropic microstructures comprised of curved surfaces were developed using a single-mask, single-etch step process. Our experimental model included HS68 normal human fibroblasts, MCF10A normal human breast epithelial cells and MDA-MB-231 metastatic human breast cancer cells. Primary findings on the first generation of silicon substrates demonstrated a distinct adhesion and growth behavior in HS68 and MDA-MB-231 cells. MDA-MB-231 cells deformed while the fibroblasts stretched and elongated their cytoskeleton on the curved surfaces. Unlike fibroblasts, MDA-MB-231 cells mainly trapped and localized inside the deep microchambers. Detailed investigations on cytoskeletal organization, adhesion pattern and morphology of the cells on the second generation of the silicon substrates demonstrated that cytoskeletal prestress and microtubules organization in HS68 cells, cell-cell junction and cell-substrate adhesion strength in MCF10A cells, and deformability of MDA-MB-231 cells (obtained by using AFM technique) affect their behavior inside the etched cavities. Treatment of MDA-MB-231 cells with experimental breast cancer drug, SAHA, on the second generation of substrates, significantly altered the cells morphology, cytoarchitecture and adhesion pattern inside the 3-D microstructures. Third generation of silicon substrates was developed for comprehensive analysis on behavior of MDA-MB-231 and MCF10A cells in a co-culture system in response to SAHA drug. Formation of colonies of both cell types was evident inside the cavities within a few hours after seeding the cells on the chips. SAHA selectively altered the morphology and cytoarchitecture in MDA-MB-231 cells. Most importantly, the majority of MDA-MB-231 cells stretched inside the etched cavities, while the adhesion pattern of MCF10A cells remained unaltered. In the last part of this dissertation, using AFM analysis, we showed that the growth medium composition has a pronounced effect on cell elasticity. Our findings demonstrated that the proposed isotropic silicon microstructures have potential applications in development of biosensor platforms for cell segregation as well as conducting fundamental biological studies. / Ph. D.

Natural polymers from renewable resources have attracted increasing interest as candidates for renewable energy and functional materials. In this work, the interactions between natural polymer thin films and biomacromolecules were studied via surface analysis techniques, such as a quartz crystal microbalance with dissipation monitoring (QCM-D), surface plasmon resonance (SPR) and atomic force microscopy (AFM). Chitinase activity on regenerated chitin (RChitin) films was studied by QCM-D and AFM. The optimal temperature and pH for chitinase activity on surfaces determined by QCM-D and AFM were consistent with bulk solution studies in the literature. Results from QCM-D also indicated that chitinase showed higher activity on fully acetylated chitin than highly deacetylated chitosan. Nanocrystalline chitin (Chitin NC) thin films were prepared by spincoating a nanocrystalline chitin colloidal suspension onto solid surfaces. Solvent exchange experiments via QCM-D with H2O/D2O revealed that Chitin NC films had more water than RChitin films of similar thickness. Results from QCM-D demonstrated that Chitin NC films had high bovine serum albumin loading capacity, and chitinase not only degraded, but also caused swelling of the chitin nanocrystals. Adsorption of human serum albumin (HSA) and fibrinogen (HFN) onto bare gold, regenerated cellulose (RC) and RChitin thin films was studied by SPR and QCM-D. Studies by SPR indicated that HSA and HFN formed close-packed monolayers on gold surfaces and sub-monolayers on polysaccharide surfaces, and the adsorption affinity of HSA for polysaccharide surfaces was greater than that of HFN. Results from QCM-D and SPR showed that the protein layers on polysaccharide surfaces had more associated water than proteins on gold surfaces. The dehydrogenative polymerization of monolignols catalyzed by physically immobilized horseradish peroxidase was investigated using QCM-D and AFM. Results from QCM-D and AFM showed that coniferyl and p-coumaryl alcohol underwent polymerization directly, whereas sinapyl alcohol required the addition of a nucleophile for polymerization. Studies by QCM-D and AFM also indicated that the surface-initiated polymerization was greatly affected by the support surface, monolignol concentration, hydrogen peroxide concentration and temperature. Thin films of dehydrogenative polymer (DHP), kraft (KL), organosolv (OL) and milled wood (MWL) lignins were used to study the enzymatic degradation of lignin mediated by lignin peroxidase (LiP) and manganese peroxidase (MnP). Results from QCM-D showed that the initial rates for degradation catalyzed by LiP increased in the order: KL < OL < MWL < guaiacyl DHP (G-DHP) < p-hydroxyphenyl DHP (H-DHP). In contrast, manganese peroxidase only degraded DHP films with a faster initial rate for G-DHP than H-DHP. Adsorption of hemicelluloses onto KL, OL and MWL thin films was studied by QCM-D and SPR. Results from QCM-D showed that hemicelluloses with different structures displayed very different adsorption behavior. Adsorption isotherms from QCM-D and SPR indicated that xyloglucan possessed stronger affinity for KL and OL films than MWL films. Data from QCM-D and SPR revealed that xyloglucan formed less hydrated layers on lignin surfaces compared to RC surfaces, and the adsorbed xyloglucan layers on different lignin films had similar percentages of coupled water. / Ph. D.

Cancer progression is accompanied with alterations in the cell biomechanical phenotype, including changes in cell structure, morphology, and responses to microenvironmental stress. These alterations result in an increased deformability of transformed cells and reduced resistance to mechanical stimuli, enabling motility and invasion. Therefore, single cell biomechanical properties could be served as a powerful label-free biomarker for effective characterization and early detection of single cancer cells. Advances and innovations in microsystems and nanotechnology have facilitated interrogation of the biomechanical properties of single cells to predict their tumorigenicity, metastatic potential, and health state. This dissertation utilized Atomic Force Microscopy (AFM) for the cell biomechanical phenotyping for cancer diagnosis and early detection, efficacy screening of potential chemotherapeutic agents, and also cancer stem-like/tumor initiating cells (CSC/TICs) characterization as the critical topics received intensive attention in the search for effective cancer treatment. Our findings demonstrated the capability of exogenous sphingosine to revert the aberrant biomechanics of aggressive cells and showed a unique, mechanically homogeneous, and extremely soft characteristic of CSC/TICs, suitable for their targeted isolation. To make full use of cell biomechanical cues, this dissertation also considered the application of nonlinear viscoelastic models such as Fractional Zener and Generalized Maxwell models for the naturally complex, heterogeneous, and nonlinear structure of living cells. The emerging need for a high-throughput clinically relevant alternative for evaluating biomechanics of individual cells led us to the development of a microfluidic system. Therefore, a high-throughput, label-free, automated microfluidic chip was developed to investigate the biophysical (biomechanical-bioelectrical) markers of normal and malignant cells. Most importantly, this dissertation also explored the biomechanical response of cells upon a dynamic loading instead of a typical transient stress. Notably, metastatic and non-metastatic cells subjected to a pulsed stress regimen exerted by AFM exhibited distinct biomechanical responses. While non-metastatic cells showed an increase in their resistance against deformation and resulted in strain-stiffening behavior, metastatic cells responded by losing their resistance and yielded slight strain-softening. Ultimately, a second generation microfluidic chip called an iterative mechanical characteristics (iMECH) analyzer consisting of a series of constriction channels for simulating the dynamic stress paradigm was developed which could reproduce the same stiffening/softening trends of non-metastatic and metastatic cells, respectively. Therefore, for the first time, the use of dynamic loading paradigm to evaluate cell biomechanical responses was used as a new signature to predict malignancy or normalcy at a single-cell level with a high (~95%) confidence level. / Ph. D.

In dieser Arbeit werden systematisch die bisher unerforschten grundlegenden physikalischen und chemischen Eigenschaften von DNA-Origami untersucht, die die Stabilität dieser aus doppelsträngiger DNA aufgebauten nanoskopischen Suprastrukturen bestimmen. In Analogie zu den zahlreichen Studien, die sich mit der Stabilität von Proteinen durch kontrollierte Denaturierung beschäftigen, spielen auch in dieser Arbeit die Denaturierungsbedingungen eine zentrale Rolle. Unter Verwendung von Guanidinium (Gdm+) als teilweise DNA-stabilisierendes, aber auch potentiell denaturierendes Kation steht dessen Wirkung auf DNA-Origami-Dreiecke im Mittelpunkt der Untersuchungen, wobei insbesondere die unerwartete Modulation der nanoskopischen Schädigung von DNA-Origami durch die begleitenden Gegenanionen zu Gdm+ im Vordergrund steht. Die Experimente zielen darauf ab, atomistische, molekulare, nanoskopische und thermodynamische Eigenschaften von DNA-Origami zu korrelieren und zu klären, wie diese vom Design des DNA-Origami selbst abhängen können. Die Ergebnisse zeigen einen unerwarteten Zusammenhang zwischen den spezifischen Gegenanionen des Denaturierungsmittels und der Stabilität der DNA-Origami-Dreiecke: Sulfat wirkt stabilisierend, während Chlorid die Superstruktur bereits unterhalb der globalen Schmelztemperatur destabilisiert. Statistische Analysen von Rasterkraftmikroskop (AFM)-Bildern und Zirkulardichroismus (CD)-Spektren zeigen Strukturübergänge auf nano-skopischer bzw. molekularer Ebene. Werden diese Techniken mit thermischer Denaturierung in Gegenwart von schwacher bis starker chemischer Denaturierung kombiniert, so zeigt sich, dass Änderungen der Wärmekapazität (ΔCp) während der strukturellen Veränderungen der DNA-Originale eine Schlüsselrolle bei der Bestimmung ihrer Empfindlichkeit gegenüber Temperatur und Denaturierungsmitteln spielen. Die Daten deuten darauf hin, dass Wasser auf apolaren DNA-Origami-Oberflächen der molekulare Ursprung der abgeleiteten Wärme-kapazitätsänderungen ist. Diese Hypothese wird durch Molekulardynamik-Simulationen (MD) unterstützt, die die Modulation von ΔCp durch die Hydratationshüllen der Anionen zeigen. Ihr unterschiedliches Potential, stabile Ionenpaare mit Gdm+ in konzentrierten Salzlösungen zu bilden, kann die experimentell beobachteten Variationen der strukturellen Stabilität erklären. Die Kopplung von strukturellen Übergängen an ΔCp wird somit als Schlüsselfaktor für die Destabilisierung von DNA-Origami sowohl bei höheren als auch bei niedrigeren Temperaturen identifiziert. Darüber hinaus weisen DNA-Origami nicht nur diese Eigenschaft auf, sondern ermöglichen auch die Beobachtung von kalten Denaturierungsprozessen auf nanoskopischer Ebene, bei denen kälteinduzierte Spannungen innerhalb der Superstruktur bei einem Bruch an vorherbestimmten lokalen Stellen freigesetzt werden, die in AFM-Bildern sichtbar sind. Dies ist die erste Beobachtung der kälteinduzierten Denaturierung von Nukleinsäuren bei Temperaturen über 0 °C sowie von DNA-basierten Superstrukturen. In dieser Arbeit wird die strukturelle Stabilität von sechs verschiedenen 2D- und 3D-DNA-Origami-Nanostrukturen in unterschiedlichen chemischen Umgebungen untersucht. Drei chaotrope Salze - Guanidiniumsulfat (Gdm2SO4), Guanidiniumchlorid (GdmCl) und Tetrapropylammoniumchlorid (TPACl) - werden als Denaturierungsmittel verwendet. Mittels Rasterkraftmikroskopie wird die Integrität der Nanostrukturen quantifiziert, wobei sich Gdm2SO4 als das schwächste und TPACl als das stärkste Denaturierungsmittel für DNA-Origami erweist, was sich auch in den Schmelztemperaturen widerspiegelt. Die Abhängigkeit der DNA-Origami-Stabilität von der Superstruktur wird besonders bei 3D-Nanostrukturen deutlich. Hier zeigen mechanisch flexible Designs sowohl in GdmCl als auch in TPACl eine höhere Stabilität als ihre starren Gegenstücke. Die Abhängigkeit der DNA-Origami-Stabilität von der Superstruktur wird besonders in 3D-Nanostrukturen deutlich, in denen mechanisch flexible Strukturen sowohl in GdmCl als auch in TPACl eine höhere Stabilität aufweisen als ihre steifen Gegenstücke. Dies begünstigt die Bildung von intramolekularen Verformungen, die sich entweder in 'weichen' Architekturen über die gesamte Superstruktur verteilen oder in ansonsten 'steifen' Strukturen in den weniger stabilen Regionen konzentrieren.:Table of contents Questions addressed in this thesis .................................................................................... I Abstract ................................................................................................................................ I Englisch .................................................................................................................................................. I Deutsch .................................................................................................................................................. II Acronyms ........................................................................................................................... III Substances .......................................................................................................................................... IV Physical and Chemical abbreviations ............................................................................................... IV Mathematical abbreviations ................................................................................................................ V 1 Introduction ................................................................................................................. 1 1.1 Deoxyribonucleic acid ................................................................................................................. 1 1.1.1 The structure of DNA ............................................................................................................ 1 1.1.2 Hydrogen bonds .................................................................................................................... 1 1.1.3 Base stacking ........................................................................................................................ 2 1.1.4 Water DNA interactions: A complex dance of stability and dynamics .................................. 5 1.1.5 The effect of ionic strength on DNA conformation ................................................................ 9 1.1.6 Conformational changes ....................................................................................................... 9 1.1.7 Forms of DNA ..................................................................................................................... 10 1.1.8 The role of apolar groups in DNA unfolding ........................................................................ 13 1.1.9 Energetics of DNA structural transitions ............................................................................. 14 1.1.10 Melting temperature ............................................................................................................ 15 1.2 Hofmeister series ....................................................................................................................... 17 1.2.1 Probing the Hofmeister series: Salt effects biomolecules ................................................... 17 1.2.2 Specific ion effects in electrolyte solutions .......................................................................... 19 1.3 DNA nanostructures................................................................................................................... 20 1.3.1 DNA origami ........................................................................................................................ 22 1.3.2 Challenges in DNA origami stability .................................................................................... 26 1.3.3 DNA origami in single molecule studies .............................................................................. 27 1.4 Circular dichroism ...................................................................................................................... 28 1.4.1 Circular dichroism spectroscopy for analyzing DNA conformations ................................... 30 1.4.2 Wavelength-dependent spectroscopic signatures of DNA conformation ........................... 32 1.5 Atomic force microscopy .......................................................................................................... 34 1.6 2D correlation spectroscopy ..................................................................................................... 36 1.6.1 2D correlation spectroscopy: Synchronous and asynchronous spectra analysis ............... 39 1.6.2 Perturbation-correlation moving-window 2D correlation spectroscopy ............................... 40 1.7 Multivariate analysis of spectral data using PCA and ITTFA ................................................ 41 1.8 Cold denaturation ....................................................................................................................... 42 2 Results and Discussion ............................................................................................ 44 2.1 Cold denaturation of the Rothemund DNA origami triangle .................................................. 45 2.2 Heat denaturation of the Rothemund DNA origami triangle .................................................. 50 2.2.1 Investigations by atomic force microscopy ......................................................................... 51 2.2.2 Circular dichroism spectroscopy and thermodynamic modelling ........................................ 56 2.2.3 Divergent effects of Cl- and SO42- on DNA origami stability ................................................ 62 2.3 Magnesium concentration modulation of DNA Origami heat denaturation ......................... 65 2.4 Assessing DNA origami stability in different chaotropic environments .............................. 66 2.4.1 DNA origami integrity influenced by Gdm2SO4 ................................................................... 67 2.4.2 DNA origami integrity influenced by GdmCl ........................................................................ 73 2.4.3 DNA origami integrity influenced by TPACl ........................................................................ 75 2.4.4 Quantitative comparison ..................................................................................................... 77 3 Critics ......................................................................................................................... 78 4 Conclusion ................................................................................................................. 79 5 Outlook ...................................................................................................................... 81 6 Material and Methods ................................................................................................ 82 6.1 DNA origami synthesis .............................................................................................................. 82 6.2 Sample preparation and AFM imaging ..................................................................................... 82 6.2.1 Anion-specific structure and stability of guanidinium-bound DNA origami & Cold denaturation of DNA origami nanostructures ...................................................................... 82 6.2.2 Superstructure-dependent stability of DNA origami nanostructures in the presence of chaotropic denaturants ........................................................................................................ 83 6.2.3 Cold denaturation of DNA origami nanostructures ............................................................. 83 6.3 CD spectroscopy and analysis ................................................................................................. 84 6.3.1 Anion-specific structure and stability of guanidinium-bound DNA origami ......................... 84 6.3.2 Pre-treatment of the CD data and calculation of melting temperatures .............................. 84 6.3.3 Cold denaturation of DNA origami nanostructures ............................................................. 84 6.3.4 Superstructure-dependent stability of DNA origami nanostructures in the presence of chaotropic denaturants ........................................................................................................ 84 6.4 Principal component analysis and iterative target test factor analysis ............................... 85 6.5 Thermodynamic modelling ........................................................................................................ 85 6.6 Molecular dynamics modelling ................................................................................................. 85 Appendix ........................................................................................................................... 88 Acknowledgment ............................................................................................................ 100 Bibliography .................................................................................................................... 101 List of Figures ................................................................................................................. 116 List of Tables ................................................................................................................... 118 Declaration of independence – Selbstständigkeitserklärung ...................................... 119 / This thesis undertakes the systematic study of hitherto unexplored fundamental physical and chemical properties of DNA origami that determine the stability of these designed nanoscopic superstructural assemblies of double-stranded DNA. In analogy to the vast number of studies addressing protein stability by controlled denaturation, denaturing conditions play a central role in this thesis as well. Using guanidinium (Gdm+) as a partly DNA-stabilizing but also potentially denaturing cation, its effect on DNA origami triangles is central to the study which particularly addressed the unexpected modulation of nanoscopic damage of DNA origami by the accompanying counter-anions to Gdm+. The experiments aim at correlating atomistic, molecular, nanoscopic and thermodynamic properties of DNA origami and at elucidating how these may depend on the DNA origami design itself. The results demonstrate an unexpected relationship between the specific counter-anions of the denaturant and the stability of DNA origami triangles: sulphate exhibits stabilizing effects and chloride induces destabilization of the superstructure already below the global melting temperature. Statistical analyses of both atomic force microscopy (AFM) images and circular dichroism (CD) spectra reveal structural transitions at the nanoscopic and molecular level, respectively. Combining these techniques with thermal denaturation in the presence of mild to strong chemical denaturation, changes in heat capacity (ΔCp) during DNA origami structural changes are shown to play the key role in determining their sensitivity to temperature and denaturants. The data suggest that water at apolar DNA origami surfaces is the molecular origin of the derived heat capacity changes. This hypothesis is substantiated by Molecular Dynamics (MD) simulations which shed light on the modulation of ΔCp by the hydration shells of anions. Their different potential to form stable ion pairs with Gdm+ in concentrated salt solutions can explain the experimentally observed variations of structural stability. The coupling of structural transitions to ΔCp is thus identified as a key factor in the destabilization of DNA origami at both elevated and lowered temperatures. Furthermore, DNA origami not only exhibit this property, but also enable the observation of cold denaturation processes at the nanoscopic level, where cold-induced strain within the superstructure is released upon breakage at predisposed local sites, visible in AFM images. This is the first observation of cold-induced denaturation of nucleic acids at temperatures above 0 °C, as well of DNA-based superstructures. Extending the scope, the work evaluates the structural stability of six different 2D and 3D DNA origami nanostructures in different chemical environments. Three chaotropic salts - guanidinium sulfate (Gdm2SO4), guanidinium chloride (GdmCl), and tetrapropylammonium chloride (TPACl) - are used as denaturants. Atomic force microscopy quantifies the nanostructural integrity, revealing Gdm2SO4 as the weakest and TPACl as the strongest DNA origami denaturant, which is also reflected in the melting temperatures. The dependence of DNA origami stability on its superstructure is particularly evident in 3D nanostructures, where mechanically flexible designs exhibit higher stability in both GdmCl and TPACl than rigid counterparts. This supports the buildup of intramolecular strain, which becomes either partitioned among the entire superstructure in “soft” architectures or accumulates at the least stable regions in otherwise “rigid” designs.:Table of contents Questions addressed in this thesis .................................................................................... I Abstract ................................................................................................................................ I Englisch .................................................................................................................................................. I Deutsch .................................................................................................................................................. II Acronyms ........................................................................................................................... III Substances .......................................................................................................................................... IV Physical and Chemical abbreviations ............................................................................................... IV Mathematical abbreviations ................................................................................................................ V 1 Introduction ................................................................................................................. 1 1.1 Deoxyribonucleic acid ................................................................................................................. 1 1.1.1 The structure of DNA ............................................................................................................ 1 1.1.2 Hydrogen bonds .................................................................................................................... 1 1.1.3 Base stacking ........................................................................................................................ 2 1.1.4 Water DNA interactions: A complex dance of stability and dynamics .................................. 5 1.1.5 The effect of ionic strength on DNA conformation ................................................................ 9 1.1.6 Conformational changes ....................................................................................................... 9 1.1.7 Forms of DNA ..................................................................................................................... 10 1.1.8 The role of apolar groups in DNA unfolding ........................................................................ 13 1.1.9 Energetics of DNA structural transitions ............................................................................. 14 1.1.10 Melting temperature ............................................................................................................ 15 1.2 Hofmeister series ....................................................................................................................... 17 1.2.1 Probing the Hofmeister series: Salt effects biomolecules ................................................... 17 1.2.2 Specific ion effects in electrolyte solutions .......................................................................... 19 1.3 DNA nanostructures................................................................................................................... 20 1.3.1 DNA origami ........................................................................................................................ 22 1.3.2 Challenges in DNA origami stability .................................................................................... 26 1.3.3 DNA origami in single molecule studies .............................................................................. 27 1.4 Circular dichroism ...................................................................................................................... 28 1.4.1 Circular dichroism spectroscopy for analyzing DNA conformations ................................... 30 1.4.2 Wavelength-dependent spectroscopic signatures of DNA conformation ........................... 32 1.5 Atomic force microscopy .......................................................................................................... 34 1.6 2D correlation spectroscopy ..................................................................................................... 36 1.6.1 2D correlation spectroscopy: Synchronous and asynchronous spectra analysis ............... 39 1.6.2 Perturbation-correlation moving-window 2D correlation spectroscopy ............................... 40 1.7 Multivariate analysis of spectral data using PCA and ITTFA ................................................ 41 1.8 Cold denaturation ....................................................................................................................... 42 2 Results and Discussion ............................................................................................ 44 2.1 Cold denaturation of the Rothemund DNA origami triangle .................................................. 45 2.2 Heat denaturation of the Rothemund DNA origami triangle .................................................. 50 2.2.1 Investigations by atomic force microscopy ......................................................................... 51 2.2.2 Circular dichroism spectroscopy and thermodynamic modelling ........................................ 56 2.2.3 Divergent effects of Cl- and SO42- on DNA origami stability ................................................ 62 2.3 Magnesium concentration modulation of DNA Origami heat denaturation ......................... 65 2.4 Assessing DNA origami stability in different chaotropic environments .............................. 66 2.4.1 DNA origami integrity influenced by Gdm2SO4 ................................................................... 67 2.4.2 DNA origami integrity influenced by GdmCl ........................................................................ 73 2.4.3 DNA origami integrity influenced by TPACl ........................................................................ 75 2.4.4 Quantitative comparison ..................................................................................................... 77 3 Critics ......................................................................................................................... 78 4 Conclusion ................................................................................................................. 79 5 Outlook ...................................................................................................................... 81 6 Material and Methods ................................................................................................ 82 6.1 DNA origami synthesis .............................................................................................................. 82 6.2 Sample preparation and AFM imaging ..................................................................................... 82 6.2.1 Anion-specific structure and stability of guanidinium-bound DNA origami & Cold denaturation of DNA origami nanostructures ...................................................................... 82 6.2.2 Superstructure-dependent stability of DNA origami nanostructures in the presence of chaotropic denaturants ........................................................................................................ 83 6.2.3 Cold denaturation of DNA origami nanostructures ............................................................. 83 6.3 CD spectroscopy and analysis ................................................................................................. 84 6.3.1 Anion-specific structure and stability of guanidinium-bound DNA origami ......................... 84 6.3.2 Pre-treatment of the CD data and calculation of melting temperatures .............................. 84 6.3.3 Cold denaturation of DNA origami nanostructures ............................................................. 84 6.3.4 Superstructure-dependent stability of DNA origami nanostructures in the presence of chaotropic denaturants ........................................................................................................ 84 6.4 Principal component analysis and iterative target test factor analysis ............................... 85 6.5 Thermodynamic modelling ........................................................................................................ 85 6.6 Molecular dynamics modelling ................................................................................................. 85 Appendix ........................................................................................................................... 88 Acknowledgment ............................................................................................................ 100 Bibliography .................................................................................................................... 101 List of Figures ................................................................................................................. 116 List of Tables ................................................................................................................... 118 Declaration of independence – Selbstständigkeitserklärung ...................................... 119

Un microscope à force atomique (AFM) est utilisé pour caractériser des matériaux isolant ou semi-conducteur avec une résolution pouvant atteindre l'échelle atomique. Ce microscope est constitué d'un capteur de force couplé à une électronique de contrôle pour pouvoir correctement caractériser ces matériaux. Parmi les différents modes (statique et dynamique), nous nous focalisons essentiellement sur le mode dynamique et plus particulièrement sur le fonctionnement sans contact à modulation de fréquence (FM-AFM). Dans ce mode, le capteur de force est maintenu comme un oscillateur harmonique par le système d'asservissement. Le projet ANR Pnano2008 intitulé : ”Cantilevers en carbure de silicium à piézorésistivité métallique pour AFM dynamique à très haute fréquence" a pour objectif d'augmenter significativement les performances d'un FM-AFM en développant un nouveau capteur de force très haute fréquence. Le but est d'augmenter la sensibilité du capteur et de diminuer le temps nécessaire à l'obtention d'une image de la surface du matériau. Le système de contrôle associé doit être capable de détecter des variations de fréquence de 100mHz pour une fréquence de résonance de 50MHz. Etant donné que les systèmes présents dans l'état de l'art ne permettent pas d'atteindre ces performances, l'objectif de cette thèse fut de développer un nouveau système de contrôle. Celui-ci est entièrement numérique et il est implémenté sur une carte de prototypage basée sur un FPGA. Dans ce mémoire, nous présentons le fonctionnement global du système ainsi que ses caractéristiques principales. Elles portent sur la détection de l'écart de fréquence de résonance du capteur de force. / An atomic force microscope (AFM) is used to characterize insulating materials or semiconductors with a resolution up to the atomic length scale. The microscope includes a force sensor linked to a control electronic in order to properly characterize these materials. Among the various modes (static and dynamic), we focus mainly on the dynamic mode and especially on the frequency modulation mode (FM-AFM). In this mode, the force sensor is maintained as a harmonic oscillator by the servo system. The research project ANR Pnano2008 entitled: "metal piezoresistivity silicon carbide cantilever for very high frequency dynamic AFM" aims to significantly increase the performance of a FM-AFM by developing new very high frequency force sensors. The goal is to increase the sensitivity of the sensor and to decrease the time necessary to obtain topography images of the material. The control system of this new sensor must be able to detect frequency variations as small as 100mHz for cantilevers with resonance frequencies up to 50MHz. Since the state-of-the-art systems doe not present these performances, the objective of this thesis was to develop a new control system. It is fully digital and it is implemented on a FPGA based prototyping board. In this report, we present the system overall functioning and its main features which are related to the cantilever resonant frequency detection. This detection is managed by a phase locked loop (PLL) which is the key element of the system.

La miniaturisation de la structure de transistor MOS a conduit à l'amincissement de l’oxyde de grille. Ainsi, la dégradation et le claquage sous contrainte électrique est devenu l'un des problèmes de fiabilité les plus importants des couches minces d'oxydes. L'utilisation de techniques de caractérisation permettant de mesurer les courants de fuite avec une résolution spatiale nanométrique a montré que le phénomène de claquage des oxydes est un phénomène très localisé. Le diamètre des «points chauds», des endroits où le courant de fuite est très élevé pour une tension appliquée continue, peut-être de quelques nanomètres uniquement. Ceci illustre pourquoi les méthodes de caractérisation avec une résolution spatiale à l’échelle nanométrique peuvent fournir des informations supplémentaires par rapport à la caractérisation classique macroscopique. Il y a deux instruments, dérivés de la microscopie à force atomique (AFM) qui peuvent être utilisés pour faire ce travail, soit le Tunneling Atomic Force Microscope (TUNA) ou le Conductive Atomic Force Microscope (C-AFM). Le mode TUNA qui est utilisé dans notre travail est capable de mesurer des courants très faibles variant entre 60 fA et 100 pA. Notre travail peut être divisé en deux thèmes principaux: - La caractérisation électrique des couches minces d'oxydes high-k (LaAlO3 et Gd2O3) à l'échelle nanométrique en utilisant le Dimension Veeco 3100 où nous avons montré que la différence de leurs techniques d'élaboration influe largement sur le comportement électrique de ces oxydes. - Les caractérisations électriques et physiques à l’échelle nanométrique des couches minces d’oxydes thermiques SiO2 sous différentes atmosphères, c.à.d. dans l'air et sous vide (≈ 10-6 mbar) en utilisant le microscope Veeco E-Scope. L'influence de l’atmosphère a été bien étudiée, où nous avons montré que les phénomènes de claquage des couches minces d'oxydes peuvent être fortement réduits sous vide surtout en l'absence du ménisque d'eau sur la surface de l'oxyde pendant les expériences. En utilisant les plusieurs modes de l'AFM, il a été démontré que l'existence de bosses anormales (hillocks) sur la surface de l'oxyde après l'application d'une tension électrique est une combinaison de deux phénomènes: la modification morphologique réelle de la surface de l'oxyde et la force électrostatique entre les charges piégées dans le volume de l'oxyde et la pointe de l'AFM. Selon les images du courant obtenues par AFM en mode TUNA, deux phénomènes physiques pour la création de ces hillocks ont été proposés: le premier est l'effet électro-thermique et la seconde est l'oxydation du substrat Si à l’interface Si/oxyde. / Miniaturization of the MOS transistor structure has led to the high thinning of the gate oxide. Hence, degradation and breakdown under electrical stress became one of the important reliability concerns of thin oxide films. The use of characterization techniques allowing to measure leakage currents with a nanometric spatial resolution has shown that breakdown phenomenon of oxides is a highly localized phenomenon. So called “hot spots”, places where the leakage current is very high for a given applied continuous voltage, can be several nanometers wide only. This illustrates why nanometric characterization methods with a nanometer range spatial resolution provide additional information compared to the classical macroscopic characterization. There are two instruments that can be used to do this job, either the Tunneling Atomic Force Microscope (TUNA) or the Conductive Atomic Force Microscope (C-AFM). TUNA which is used in our work is capable to measure very low currents ranging between 60 fA and 100 pA. Our work can be divided into two principle topics: - Electrical characterization of thin high-k oxides (LaAlO3 and Gd2O3) at the nano-scale using the Veeco Dimension 3100 where we have shown that the difference in their elaboration techniques largely influence the electrical behavior of these oxides. - Nano-scale electrical and physical characterization of thin SiO2 thermal oxides in different surrounding ambient, that is in air and under vacuum (≈ 10-6 mbar) using the Veeco E-scope microscope. The influence of the experiment surrounding ambient has been well studied where we have shown that the breakdown phenomena of thin oxide films can be highly reduced under vacuum especially in the absence of the water meniscus on the oxide’s surface under study. Using different AFM modes, it was demonstrated that the existence of the well-known hillock (protrusions) on the oxide’s surface after the application of an electrical stress is a combination of two phenomena: the real morphological modification of the oxide’s surface and the electrostatic force between the trapped charges in the oxide’s volume and the AFM tip. Depending on the current images obtained by TUNA AFM mode, two physical phenomena for the creation of these hillocks have been proposed: the first is the electro-thermal effect and the second is the oxidation of the Si substrate at the Si/oxide interface.

Recently, remarkable advances have been made in the understanding of micro/nanoscale energy transport, opening new opportunities in various areas such as thermal management, data storage, and energy conversion. This dissertation focuses on thermally-sensed nanotopography using a heated silicon microcantilever and near-field thermophotovoltaic (TPV) energy conversion system. A heated microcantilever is a functionalized atomic force microscope (AFM) cantilever that has a small resistive heater integrated at the free end. Besides its capability of increasing the heater temperature over 1,000 K, the resistance of a heated cantilever is a very sensitive function of temperature, suggesting that the heated cantilever can be used as a highly sensitive thermal metrology tool. The first part of the dissertation discusses the thermal characterization of the heated microcantilever for its usage as a thermal sensor in various conditions. Particularly, the use of heated cantilevers for tapping-mode topography imaging will be presented, along with the recent experimental results on the thermal interaction between the cantilever and substrate. In the second part of the dissertation, the so-called near-field TPV device is introduced. This new type of energy conversion system utilizes the significant enhancement of radiative energy transport due to photon tunneling and surface polaritons. Investigation of surface and bulk polaritons in a multilayered structure reveals that radiative properties are significantly affected by polariton excitations. The dissertation then addresses the rigorous performance analysis of the near-field TPV system and a novel design of a near-field TPV device.

Ziel dieser Arbeit ist es, eine Verbindung zwischen der Mikrostruktur teilkristalliner Polymere und derer mechanischen Eigenschaften auf der Mikro- und Nanometerskala aufzubauen. Dazu wurden Methoden der Rasterkraftmikroskopie verwendet um sowohl orts- als auch zeitaufgelöst Kristallisations-, Deformations- und Diffusionsprozesse in der Mikrostruktur von elastomerem Polypropylen (ePP) abzubilden. Die mechanischen Eigenschaften wurden simultan mit Mikrozugversuchen bestimmt. So konnte beispielsweise ein Zusammenhang zwischen abnehmender Kristall-Kristall-Distanz und einem Ansteigen des Elastizitätsmoduls während der Kristallisation nachgewiesen werden. Weiterhin war es möglich die Veränderung der nano-mechanischen Eigenschaften während der Kristallisation einzelner kristalliner Lamellen in deren direkter Umgebung mit MUSIC-mode Rasterkraftmikroskopie zu untersuchen. Laterale Querexpansion (auxetisches Verhalten) konnte bei uniaxialen Zugversuchen für die Kreuzschraffur-Struktur elastomeren Polypropylens auf der Größenskala einiger Mikrometer nachgewiesen werden. Zusätzlich wurde eine Orientierungsabhängigkeit dieses Effekts beobachtet. Außerdem wurde die Diffusion einzelner Kristalle in der Mikrostruktur von ePP beobachtet. Die Heterogenität dieser Diffusion lässt auf eine kristallin-amorph Grenzschicht um alle Kristalle schließen.