Carnitine tailored Sensors on Surface Molecular Imprinting based on Graphene layers

III Jornadas de Electroquímica e Inovação (Electroquímica e Nanomateriais), na Universidade de Trás-os-Montes e Alto Douro, Vila Real, 16 a 17 de Setembro de 2013

Host-Tailored Sensors for Carnitine Potentiometric Measurements based on Surface Molecular Imprinting

Graduate Student Symposium on Molecular Imprinting 2013, na Queen’s University, Belfast, United Kingdom, 15 a 17 de Agosto de 2013

Multi-stacks of epitaxial GeSn self-assembled dots in Si: Structural analysis

We report on the growth and structural and morphologic characterization of stacked layers of self-assembled GeSn dots grown on Si (100) substrates by molecular beam epitaxy at low substrate temperature T = 350 °C. Samples consist of layers (from 1 up to 10) of Ge0.96Sn0.04 self-assembled dots separated by Si spacer layers, 10 nm thick. Their structural analysis was performed based on transmission electron microscopy, atomic force microscopy and Raman scattering. We found that up to 4 stacks of dots could be grown with good dot layer homogeneity, making the GeSn dots interesting candidates for optoelectronic device applications.

Effects of physiological self-crowding of DNA on shape and biological properties of DNA molecules with various levels of supercoiling.

DNA in bacterial chromosomes and bacterial plasmids is supercoiled. DNA supercoiling is essential for DNA replication and gene regulation. However, the density of supercoiling in vivo is circa twice smaller than in deproteinized DNA molecules isolated from bacteria. What are then the specific advantages of reduced supercoiling density that is maintained in vivo? Using Brownian dynamics simulations and atomic force microscopy we show here that thanks to physiological DNA-DNA crowding DNA molecules with reduced supercoiling density are still sufficiently supercoiled to stimulate interaction between cis-regulatory elements. On the other hand, weak supercoiling permits DNA molecules to modulate their overall shape in response to physiological changes in DNA crowding. This plasticity of DNA shapes may have regulatory role and be important for the postreplicative spontaneous segregation of bacterial chromosomes.

Cysteine-rich Domain 1 of CD40 Mediates Receptor Self-assembly.

The activation of CD40 on B cells, macrophages, and dendritic cells by its ligand CD154 (CD40L) is essential for the development of humoral and cellular immune responses. CD40L and other TNF superfamily ligands are noncovalent homotrimers, but the form under which CD40 exists in the absence of ligand remains to be elucidated. Here, we show that both cell surface-expressed and soluble CD40 self-assemble, most probably as noncovalent dimers. The cysteine-rich domain 1 (CRD1) of CD40 participated to dimerization and was also required for efficient receptor expression. Modelization of a CD40 dimer allowed the identification of lysine 29 in CRD1, whose mutation decreased CD40 self-interaction without affecting expression or response to ligand. When expressed alone, recombinant CD40-CRD1 bound CD40 with a KD of 0.6 μm. This molecule triggered expression of maturation markers on human dendritic cells and potentiated CD40L activity. These results suggest that CD40 self-assembly modulates signaling, possibly by maintaining the receptor in a quiescent state.

Investigation on diamond turning of silicon crystal - generation mechanism of surface cut with worn tool

This paper discusses the effect of tool wear on surface finish in single-point diamond turning of single crystal silicon. The morphology and topography of the machined surface clearly show the type of cutting edge wear reproduced onto the cutting grooves. Scanning electron microscopy is used in order to correlate the cutting edge damage and microtopography features observed through atomic force microscopy. The possible wear mechanisms affecting tool performance and surface generation during cutting are also discussed. The zero degree rake angle single point diamond tool presented small nicks on the cutting edge. The negative rake angle tools presented more a type of crater wear on the rake face. No wear was detected on flank face of the diamond tools.

Synthesis and characterization of nanoperforated metal oxide thin films

For advanced devices in the application fields of data storage, solar cell and biosensing, one of the major challenges to achieve high efficiency is the fabrication of nanopatterned metal oxide surfaces. Such surfaces often require both precise structure at the nanometer scale and controllable patterned structure at the macro scale. Nowadays, the dominating candidates to fabricate nanopatterned surfaces are the lithographic technique and block-copolymer masks, most of which are unfortunately costly and inefficient. An alternative bottom-up approach, which involves organic/inorganic self-assembly and dip-coating deposition, has been studied intensively in recent years and has proven to be an effective technique for the fabrication of nanoperforated metal oxide thin films. The overall objective of this work was to optimize the synthesis conditions of nanoperforated TiO2 (NP-TiO2) thin films, especially to be compatible with mixed metal oxide systems. Another goal was to develop fabrication and processing of NP-TiO2 thin films towards largescale production and seek new applications for solar cells and biosensing. Besides the traditional dip-coating and drop-casting methods, inkjet printing was used to prepare thin films of metal oxides, with the advantage of depositing the ink onto target areas, further enabling cost-effective fabrication of micro-patterned nanoperforated metal oxide thin films. The films were characterized by water contact angle determination, Atomic Force Microscopy, Scanning Electron Microscopy, X-ray Photoelectron Spectroscopy and Grazing Incidence XRay Diffraction. In this study, well-ordered zinc titanate nanoperforated thin films with different Zn/Ti ratios were produced successfully with zinc precursor content up to 50 mol%, and the dominating phase was Zn2Ti3O8. NP-TiO2 structures were also obtained by a cost-efficient means, namely inkjet printing, at both ambient temperature and 60 °C. To further explore new biosensing applications of nanoperforated oxide thin films, inkjet printing was used for the fabrication of both continuous and patterned polymeric films onto NP-TiO2 and perfluorinated phosphate functionalized NP-TiO2 substrates, respectively. The NP-TiO2 films can be also functionalized with a fluoroalkylsilane, resulting in hydrophobic surfaces on both titania and silica. The surface energy contrast in the nanoperforations can be tuned by irradiating the films with UV light, which provides ideal model systems for wettability studies.

Assessing the Reproducibility of Clustering of Molecular Dynamics Conformations on Self-Organizing Maps

The goal of most clustering algorithms is to find the optimal number of clusters (i.e. fewest number of clusters). However, analysis of molecular conformations of biological macromolecules obtained from computer simulations may benefit from a larger array of clusters. The Self-Organizing Map (SOM) clustering method has the advantage of generating large numbers of clusters, but often gives ambiguous results. In this work, SOMs have been shown to be reproducible when the same conformational dataset is independently clustered multiple times (~100), with the help of the Cramérs V-index (C_v). The ability of C_v to determine which SOMs are reproduced is generalizable across different SOM source codes. The conformational ensembles produced from MD (molecular dynamics) and REMD (replica exchange molecular dynamics) simulations of the penta peptide Met-enkephalin (MET) and the 34 amino acid protein human Parathyroid Hormone (hPTH) were used to evaluate SOM reproducibility. The training length for the SOM has a huge impact on the reproducibility. Analysis of MET conformational data definitively determined that toroidal SOMs cluster data better than bordered maps due to the fact that toroidal maps do not have an edge effect. For the source code from MATLAB, it was determined that the learning rate function should be LINEAR with an initial learning rate factor of 0.05 and the SOM should be trained by a sequential algorithm. The trained SOMs can be used as a supervised classification for another dataset. The toroidal 10×10 hexagonal SOMs produced from the MATLAB program for hPTH conformational data produced three sets of reproducible clusters (27%, 15%, and 13% of 100 independent runs) which find similar partitionings to those of smaller 6×6 SOMs. The χ^2 values produced as part of the C_v calculation were used to locate clusters with identical conformational memberships on independently trained SOMs, even those with different dimensions. The χ^2 values could relate the different SOM partitionings to each other.

Self-assembly of PS-PVP block copolymers and their complexes at the air/water interface

Une compréhension approfondie et un meilleur contrôle de l'auto-assemblage des copolymères diblocs (séquencés) et de leurs complexes à l'interface air/eau permettent la formation contrôlée de nanostructures dont les propriétés sont connues comme alternative à la nanolithographie. Dans cette thèse, des monocouches obtenues par les techniques de Langmuir et de Langmuir-Blodgett (LB) avec le copolymère dibloc polystyrène-poly(4-vinyl pyridine) (PS-PVP), seul ou complexé avec de petites molécules par liaison hydrogène [en particulier, le 3-n-pentadécylphénol (PDP)], ont été étudiées. Une partie importante de notre recherche a été consacrée à l'étude d'une monocouche assemblée atypique baptisée réseau de nanostries. Des monocouches LB composées de nanostries ont déjà été rapportées dans la littérature mais elles coexistent souvent avec d'autres morphologies, ce qui les rend inutilisables pour des applications potentielles. Nous avons déterminé les paramètres moléculaires et les conditions expérimentales qui contrôlent cette morphologie, la rendant très reproductible. Nous avons aussi proposé un mécanisme original pour la formation de cette morphologie. De plus, nous avons montré que l'utilisation de solvants à haut point d’ébullition, non couramment utilisés pour la préparation des films Langmuir, peut améliorer l'ordre des nanostries. En étudiant une large gamme de PS-PVP avec des rapports PS/PVP et des masses molaires différents, avec ou sans la présence de PDP, nous avons établi la dépendance des types principaux de morphologie (planaire, stries, nodules) en fonction de la composition et de la concentration des solutions. Ces observations ont mené à une discussion sur les mécanismes de formation des morphologies, incluant la cinétique, l’assemblage moléculaire et l’effet du démouillage. Nous avons aussi démontré pour la première fois que le plateau dans l'isotherme des PS-PVP/PDP avec morphologie de type nodules est relié à une transition ordre-ordre des nodules (héxagonal-tétragonal) qui se produit simultanément avec la réorientation du PDP, les deux aspects étant clairement observés par AFM. Ces études ouvrent aussi la voie à l'utilisation de films PS-PVP/PDP ultraminces comme masque. La capacité de produire des films nanostructurés bien contrôlés sur différents substrats a été démontrée et la stabilité des films a été vérifiée. Le retrait de la petite molécule des nanostructures a fait apparaître une structure interne à explorer lors d’études futures.

Molecular tectonics : supramolecular 2D nanopatterning of surfaces by self-assembly

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal

Détection de protéines par diffusion Raman exaltée par effet de pointe (TERS)

La concentration locale des messagers chimiques sécrétés par les cellules peut être mesurée afin de mieux comprendre les mécanismes moléculaires liés à diverses maladies, dont les métastases du cancer. De nouvelles techniques analytiques sont requises pour effectuer ces mesures locales de marqueurs biologiques à proximité des cellules. Ce mémoire présentera le développement d’une nouvelle technique basée sur la réponse plasmonique sur des leviers AFM, permettant d’étudier les réactions chimiques et biologiques à la surface des leviers grâce au phénomène de résonance des plasmons de surface (SPR), ainsi qu’à la diffusion Raman exaltée par effet de pointe (TERS). En effet, il est possible de localiser l’amplification du signal Raman à la pointe d’un levier AFM, tout comme le principe de la diffusion Raman exaltée par effet de surface (SERS) basée sur la diffusion de la lumière par des nanoparticules métalliques, et permettant une large amplification du signal Raman. La surface du levier est recouverte d’une nano-couche métallique d’or, suivi par des réactions biologiques pour l’immobilisation d’un récepteur moléculaire, créant ainsi un biocapteur sur la pointe du levier. Une détection secondaire utilisant des nanoparticules d’or conjuguées à un anticorps secondaire permet également une amplification du signal SPR et Raman lors de la détection d’antigène. Ce mémoire démontrera le développement et la validation de la détection de l’immunoglobuline G (IgG) sur la pointe du levier AFM.Dans des projets futurs, cette nouvelle technique d’instrumentation et d’imagerie sera optimisée grâce à la création d’un micro-détecteur protéique généralement adapté pour l’étude de la communication cellulaire. En intégrant le signal SPR à la microscopie AFM, il sera alors possible de développer des biocapteurs SPR couplés à une sonde à balayage, ce qui permettra d’effectuer une analyse topographique et de l’environnement chimique d’échantillons cellulaires en temps réel, pour la mesure des messagers moléculaires sécrétés dans la matrice extracellulaire, lors de la communication cellulaire.

Mechanisms and dynamics of protein clustering on a solid surface

A methodology for discovering the mechanisms and dynamics of protein clustering on solid surfaces is presented. In situ atomic force microscopy images are quantitatively compared to Monte Carlo simulations using cluster statistics to differentiate various models. We study lysozyme adsorption on mica as a model system and find that all surface-supported clusters are mobile, not just the monomers, with diffusion constant inversely related to cluster size. The surface monomer diffusion constant is measured to be D1∼9×10-16  cm2 s-1, such a low value being difficult to measure using other techniques.

New insights on growth mechanisms of protein clusters at surfaces: an AFM and simulation study

Despite its relevance to a wide range of technological and fundamental areas, a quantitative understanding of protein surface clustering dynamics is often lacking. In inorganic crystal growth, surface clustering of adatoms is well described by diffusion-aggregation models. In such models, the statistical properties of the aggregate arrays often reveal the molecular scale aggregation processes. We investigate the potential of these theories to reveal hitherto hidden facets of protein clustering by carrying out concomitant observations of lysozyme adsorption onto mica surfaces, using atomic force microscopy. and Monte Carlo simulations of cluster nucleation and growth. We find that lysozyme clusters diffuse across the substrate at a rate that varies inversely with size. This result suggests which molecular scale mechanisms are responsible for the mobility of the proteins on the substrate. In addition the surface diffusion coefficient of the monomer can also be extracted from the comparison between experiments and simulations. While concentrating on a model system of lysozyme-on-mica, this 'proof of concept' study successfully demonstrates the potential of our approach to understand and influence more biomedically applicable protein-substrate couples.

Influence of the solvent on the self-assembly of a modified amyloid beta peptide fragment. I. Morphological investigation

The solvent-induced transition between self-assembled structures formed by the peptide AAKLVFF is studied via electron microscopy, light scattering, and spectroscopic techniques. The peptide is based on a core fragment of the amyloid beta-peptide, KLVFF, extended by two alanine residues. AAKLVFF exhibits distinct structures of twisted fibrils in water or nanotubes in methanol. For intermediate water/methanol compositions, these structures are disrupted and replaced by wide filamentous tapes that appear to be lateral aggregates of thin protofilaments. The orientation of the beta-strands in the twisted tapes or nanotubes can be deduced from X-ray diffraction on aligned stalks, as well as FT-IR experiments in transmission compared to attenuated total reflection. Strands are aligned perpendicular to the axis of the twisted fibrils or the nanotubes. The results are interpreted in light of recent results on the effect of competitive hydrogen bonding upon self-assembly in soft materials in water/methanol mixtures.

Stepwise self-assembly of a tripeptide from molecular dimers to supramolecular beta-sheets in crystals and amyloid-like fibrils in the solid state

The terminally protected tripeptide Boc-Ala(1)-Leu(2)-Ala(3)-OMe 1 forms antiparallel hydrogen-bonded dimers of two different conformers in the asymmetric unit and the individual dimers then self-associate to form supramolecular beta-sheet structures in crystals and amyloid-like fibrils in the solid state.