Plasmonic antennas and zero mode waveguides to enhance single molecule fluorescence detection and fluorescence correlation spectroscopy towards physiological concentrations

Single-molecule approaches to biology offer a powerful new vision to elucidate the mechanisms that underpin the functioning of living cells. However, conventional optical single molecule spectroscopy techniques such as F\"orster fluorescence resonance energy transfer (FRET) or fluorescence correlation spectroscopy (FCS) are limited by diffraction to the nanomolar concentration range, far below the physiological micromolar concentration range where most biological reaction occur. To breach the diffraction limit, zero mode waveguides and plasmonic antennas exploit the surface plasmon resonances to confine and enhance light down to the nanometre scale. The ability of plasmonics to achieve extreme light concentration unlocks an enormous potential to enhance fluorescence detection, FRET and FCS. Single molecule spectroscopy techniques greatly benefit from zero mode waveguides and plasmonic antennas to enter a new dimension of molecular concentration reaching physiological conditions. The application of nano-optics to biological problems with FRET and FCS is an emerging and exciting field, and is promising to reveal new insights on biological functions and dynamics.Comment: WIREs Nanomed Nanobiotechnol 201

Current methods for characterising mixing and flow in microchannels

This article reviews existing methods for the characterisation of mixing and flow in microchannels, micromixers and microreactors. In particular, it analyses the current experimental techniques and methods available for characterising mixing and the associated phenomena in single and multiphase flow. The review shows that the majority of the experimental techniques used for characterising mixing and two-phase flow in microchannels employ optical methods, which require optical access to the flow, or off-line measurements. Indeed visual measurements are very important for the fundamental understanding of the physics of these flows and the rapid advances in optical measurement techniques, like confocal scanning laser microscopy and high resolution stereo micro particle image velocimetry, are now making full field data retrieval possible. However, integration of microchannel devices in industrial processes will require on-line measurements for process control that do not necessarily rely on optical techniques. Developments are being made in the areas of non-intrusive sensors, magnetic resonance techniques, ultrasonic spectroscopy and on-line flow through measurement cells. The advances made in these areas will certainly be of increasing interest in the future as microchannels are more frequently employed in continuous flow equipment for industrial applications

Modelling thin film growth in the Ti-Ag system

With the aim to model the surface growth of Ti-Ag system over realistic time scales, two interatomic potential mixing rules for the Ti-Ag system were first investigated based on the embedded-atom method (EAM) elemental potentials. First principles calculations were performed using SIESTA for various configurations of the Ti-Ag system to see which model best fitted the ab initio results. The results showed that the surface energies, es- pecially that of Ti, were not well fitted by either model and the surface binding energies differed from the ab initio calculations. As a result, the modified embedded-atom method (MEAM) was investigated. In contrast to the other models, surface energies for pure Ti calculated by MEAM were in good agreement with the experimental data and the ab initio results. The MEAM mixing rule was used to investigate Ag adatoms on Ti and Ti adatoms on Ag. The results showed good agreement with SIESTA after parameter optimisation. Simulations of thin film growth in the Ag-Ti system are presented using an adaptive kinetic Monte Carlo method (AKMC). For the growth of Ti on Ag (100) and Ag (111) surfaces, the Ti adatoms prefer to exchange with the original surface layer atoms creating a mixed Ag/Ti surface. On a silver substrate, up to four mixed layers need to be formed before a pure Ti layer is obtained when the deposition energy is less than 20 eV. Conversely, the simulations of Ag on the Ti (0001) plane showed that the Ag adatoms repel each other on the Ti basal plane, before a complete first layer of Ag was obtained in a face-centred cubic structure. The implementations of a super-basin method within the adaptive ki- netic Monte Carlo method has allowed the simulation of 0.4s of surface growth on the Ag substrates. This work also compared two long time scale dynamics methods, namely AKMC and Parallel Trajectory Splicing (ParSplice) simulations. For these two configurations are considered on the Ag (111) substrate. The transitions and the associated energy barriers are identical for single atom diffusion but the diffusion rates differ. In the case of an adatom on an island, a super-basin system was created. The exit transitions found by a transition search algorithm and ParSplice were again the same whilst the mean exit time differed by a factor of two due to inaccurate prefactor calculations. The distribution of basin-exit times is also examined which obeys an exponential distribution

Advances in biophysical methods for protein detection and characterisation

Proteins are the building blocs of life and mediate nearly every function in the cell. They are therefore a major and incredibly wide research topic. Their functions and malfunctions have serious impacts on a wide range of diseases. Proteins may be used as extremely versatile tools for biology such as for gene editing or biological medical products. Developing novel methods for protein detection and characterisation may thus have a tremendous impact on modern medicine and research. The present thesis discusses advances in biophysical methods for protein detection and characterisation. First, the possibility of detecting proteins label-free is addressed. A label may change the behaviour of the target protein. Two approaches are investigated: An ultraviolet light based autofluorescence microscope is described; and scattering based detection is explored by expending on the existing interferometric scattering (iSCAT) technique. An oblique illumination approach helps with increasing the contrast of the data, and a time correlation technique is used for local sizing on chip. Second, microfluidic techniques are routinely used to create protein assays. These assays minimise the amount of sample required and the absence of turbulences enable new techniques. A method to easily add nanofluidics elements to microfluidic designs is discussed. Finally, three characterisation methods are described. First, diffusional sizing uses a microfluidic chip to create a concentration gradient. The protein diffusion coefficient is extracted from the time evolution of this gradient. Second, the diffusiophoretic coefficient of the protein can be extracted by diffusiophoresis, which is the motion of proteins driven by the concentration gradient of another solute. This could be an important protein motion mechanism, as many gradients are present in cells and in living beings. Finally, the spatial propagation of the protein amyloid-beta 1-42, a protein associated to neurodegenerative disorders, is observed in a capillary

Simulating radiation damage in austenitic stainless steel and Ni-based alloys

The evolution of materials at an atomistic level may have vital consequences for the properties of materials. Therefore, modelling long time scale behaviour of defects in a material is very important, particularly for those used in nuclear power plants. The materials used in nuclear power plants should have good mechanical properties to overcome the corrosive environment and high temperature. Examples of these materials are the austenitic stainless steel and the Ni-based alloys due to their high temperature properties. Molecular Dynamics (MD) and on the fly Kinetic Monte Carlo (otf-KMC) techniques have been used to model the radiation damage in austenitic stainless steel and the Ni-based alloys. This thesis represents the main findings obtained. Three potentials were implemented and used to study radiation damage in austenitic stainless steel. Structural properties such as the elastic constants for the point defects in the pure metals were first calculated. This was followed by calculating the formation energies and migration energies of vacancy and self interstitial defects in the pure metals. Different calculations were performed using each potential on the ternary alloy (Fe with 10 at.% Ni and 20 at.% Cr) and the binary alloy (Ni with 20 at.% Cr) . For example, the segregation in these alloys was investigated using Monte Carlo simulations and results obtained for both alloys at high temperature MD. Furthermore, the vacancy formation energies were calculated for both alloys using all the potentials. Radiation damage at Grain Boundaries (GBs) in fcc Ni and a Ni-Cr binary alloy has been studied using MD and otf-KMC techniques. From the results obtained, the mobility of interstitials were found to be higher than that of vacancies and tend to move quickly to the GB. Vacancies are found to migrate to the GB if they are near otherwise they tend to form clusters in the bulk. During the simulations, interesting mechanisms were observed for the point defects migration and recombinations. Large roughening at the GB was observed, especially in the alloy system and overall the total number of defects accumulated on the GB after multiple collision cascades were relatively small. The radiation in fcc Ni resulting from low energy collision cascades was also modelled using MD and otf-KMC techniques. This part of work aimed replicating the observations seen in experiment and trying to understand them. Recombinations between vacancies and interstitials were found to happen from large distances with low barriers. Most defects produced from low energy collision cascades were found to recombine or interstitials were found to form clusters. Modelling the evolution of the vacancies shows the possibility of producing Stacking Fault Tetrahedra (SFT) which were found to dissociate at 200°C

The effect of water dynamics on conformation changes of albumin in pre-denaturation state:photon correlation spectroscopy and simulation

Water is essential for protein three-dimensional structure, conformational dynamics, and activity. Human serum albumin (HSA) is one of major blood plasma proteins, and its functioning is fundamentally determined by the dynamics of surrounding water. The goal of this study is to link the conformational dynamics of albumin to the thermal motions in water taking place in the physiological temperature range. We report the results of photon correlation spectroscopy and molecular dynamics simulations of HSA in aqueous solution. The experimental data processing produced the temperature dependence of the HSA hydrodynamic radius and its zeta potential. Molecular dynamics reproduced the results of experiments and revealed changes in the secondary structure caused by the destruction of hydrogen bonds in the macromolecule's globule

Non-equilibrium polymeric complex fluids

Complex fluids are commercially- and industrially-important materials which exhibit ordering on scales much larger than atomic. Their usage is typically in non-equilibrium conditions, however traditional methods for measuring rheology are not appropriate for measuring samples with gradients present, such as temperature and concentration. In this work a safe and easy to use optical tweezer (OT) apparatus has been developed in order to facilitate the investigation of various systems during dilution or drying. In contrast to other OT setups, this equipment is safe to use without laser goggles or interlocked rooms, yet still allows full access to the microscope. Proof-of-concept experiments are performed on aqueous poly (ethylene oxide) (PEO) solutions to demonstrate the changes in viscosity and concentration over time, and the OT is then used in a rheological investigation into a commercially-relevant wormlike micelle (WLM) system, in conjunction with Diffusing Wave Spectroscopy (DWS) and traditional bulk rheology. It is shown for the first time that equimolar (eM) SDS:CAPB WLM samples can be considered ‘model’ systems, and form close approximations of Maxwellian systems on the addition of extra salt or surfactant above 0.1eM. The effect of an uncharged polymer (PEO 4M MW) on this WLM network structure was subsequently investigated; its effects are consistent with current theories of polymer-surfactant interactions

An exploration of how a drama-based pedagogy can promote understanding of chemical concepts in 11-15 year old science students

A growing body of evidence suggests that some Science teachers use drama-based strategies in order to promote understanding of abstract scientific concepts. These strategies employ action and imagination to simulate systems and processes that are too fast, too slow, too big, too small, too expensive or too dangerous to observe in the classroom. A small group of quantitative and qualitative studies over the past thirty years has suggested that these physical simulations enable learning in secondary students, by promoting discourse and by conveying concept features through a range of sensations. The field is as yet under-theorised, consisting of single case designs and unreplicated methodologies. This multiple case study focused upon an intervention design based on a pedagogical model developed in my Masters research. This study aimed to explore the characteristics of students’ interaction and the nature of their resultant conceptions over four months. Each case focussed upon one of eight Key Stage 3 and Key Stage 4 classes across a variety of UK schools. In each, a curriculum-based particle theory topic was taught in a double-period lesson. Data included video, participant observations, and interviews with three students from each class collected at pre, post and delayed intervals. Findings suggested that the pedagogy engendered engagement and self-regulation in group model-making tasks, and supported thought experiment-type visualisations of dynamic processes. Conceptual development was found to continue up to four months after the lessons. A model of learning was developed in which social interaction and multimodal discourse promoted the association of conceptual features with affective, visual and embodied images, which supported recall, discussion and further conceptual development in the longer term

Computational Investigation of Crystalline and Amorphous Borosilicates

By utilising both Molecular Dynamics (MD) and Density Functional Theory (DFT), the project focuses on the structure and bonding in glassy and crystalline boron oxide and borates, particularly on the distribution of BO3 and BO4 groups though ring and structural analysis and on the bonding and structural properties of borates. There are two main areas of this research: the first section presents the results of the computational investigation of Boron Trioxide (B2O3) via MD. The second focuses on the alkali-infused crystalline borates to understand their network structure in detail. The MD component of this investigation concentrates on pure B2O3 including the crystalline, molten, and glassy structure. The atomic structure of this material is one that has been widely debated and was a key topic in this project. Characteristics are identified through various structural and mechanical properties and via ring analysis. The results of the initial MD simulations indicate a substantial proportion of BO3 groups present in the structure of pure B2O3 (87.5%). However, expanding the simulation cell size reveals an increase in the BO4 subgroup, indicating a possibility of an increase in boroxol ring formation, this finding demonstrates the need for large simulation cells which is now possible with contemporary computational resources. The thesis then focusses on the testing and identification of a force field to utilise in the MD studies. It explores the two available force fields, containing two- and threebody terms and shows that both can model the molten structure. However, as the Computational Investigation of Crystalline and Amorphous Borosilicates quenching of the simulation takes place, the two-body potential fails to adequately create the glassy phase; whereas the potential with the three-body term is able to achieve the glassy-B2O3 structures at a much quicker rate. Crystalline borates with a range Na2O/B2O3 ratios are next investigated using Density Functional Theory. Various crystals compositions are analysed, with a focus primarily on electronic and mechanical properties. We show that an increase in sodium content in these types of crystals has a direct effect on elasticity and mechanical strength. The results reveal interesting aspects of the bonding in these materials and give insight into the relationship between structure and composition

Modelling thin film growth over realistic time scales

Energy security and supply is a key problem for the UK in the coming years. Photovoltaics have an important role to play in this. In order for demand to be met, continued research into new materials and methods of production is necessary. By modelling deposition techniques using classical molecular dynamics (MD), an atomistic scale understanding can be obtained. Combining this with long time scale dynamics (LTSD) techniques allows us to also model diffusion and surface growth over realistic time scales. The LTSD technique applied throughout this project is an on-the-fly Kinetic Monte Carlo (otf-KMC) method, which determines diffusion pathways and barriers, in parallel, with no prior knowledge of the involved transitions. These simulation techniques allow parameters such as deposition energy, substrate bias and plasma pressure to be easily changed to gain understanding of their effects. During this project, growth via industrial scale deposition techniques has been simulated, including evaporation (thermal and electron beam), ion-beam assisted evaporation and reactive magnetron sputtering. Metal thin films, of interest due to their uses in reflectors in concentrator photovoltaics, electrical conductors in the monolithic interconnect processes and back contacts, were investigated using otf-KMC. Ag and Al film growth was simulated for around 0.3 seconds of real time. It was found that Ag has the ability to grow smooth surfaces, using several mechanisms including multiple-atom concerted motion, exchange mechanisms, and damage and repair mechanisms. Ag (111) and (100) surfaces grew dense, complete and crystalline films when sputtering was simulated, however, evaporation deposition produced incomplete layers. The inclusion of Ar in the ion-beam assisted evaporation of Ag (111) aided growth by transferring extra energy to the surface allowing increased diffusion and atomic mixing. Al (111) and (100), however, show different patterns. Growth by evaporation deposition and magnetron sputtering actually produced very similar results. The inclusion of the ion-beam assist on the (111) surface actually damaged the film, producing subsurface Ar clusters where Al atoms were displaced, creating voids throughout the film. Otf-KMC methods enabled the investigation of specific mechanisms allowing film growth and a very important transition enabling the smooth and complete Al film growth was found to be the Ehrlich-Schwoebel (ES) barrier. The ES barrier involves an atom dropping off a step edge of an island and this barrier was found to be much smaller for the Al surfaces, therefore allowing the more complete growth from both evaporation and sputtering. Metal oxides are also of great interest in the photovoltaic industry. The rutile TiO$_2$ (110) surface was investigated using single point depositions, high temperature MD and otf-KMC. Otf-KMC enabled the simulation for up to 9 seconds of real time, totally inaccessible using traditional simulation methods. Results concluded that the evaporation deposition process produced a void filled, incomplete structure, even with the use of a low energy ion-beam assist, this material is of interest for dye-sensitised solar cells where a dye is injected into the voids. Sputtering, however, produced dense and crystalline film, which is much more applicable to anti-reflective coatings where a crystalline structure is required. Mechanisms which enabled crystalline rutile to form were also investigated, highlighting Ti interstitial annealing in the presence of an O rich surface as an important rutile growth mechanism. ZnO, an inorganic compound with many uses including transparent conductive oxides, is investigated in the most stable wurtzite phase. The O-terminated (000$\bar{1}$) polar surface was used as the substrate for otf-KMC growth simulations, where around 1 second of real time was simulated. Evaporation deposition of a stoichiometric distribution of deposition species was found to produce the best quality film, however, a phase boundary was observed where an area of zinc blende forms within the wurtzite. Sputtering resulted in a denser, more complete and crystalline structure due to the higher deposition energy of arriving species, similar to the TiO$_2$ results. Post-annealing at 770K did not allow complete recrystallisation, resulting in films with stacking faults where monolayers formed in the zinc blende phase. Annealing at 920K, however, in some cases enabled the complete recrystallisation of films back into the wurtzite structure. Although, the higher annealing temperature did not always enable recrystallisation and in some cases both wurtzite and zinc blende phases existed in the same layer, resulting in a phase boundary. An important mechanism for the nucleation of ZnO growth was found to be the formation and vibration of Zn$_x$O$_y$ strings on the surface, which after hundreds of milliseconds formed the desired hexagonal structure. Combining MD and otf-KMC enabled the simulation of systems over very large time scales which were previously totally inaccessible. Key mechanisms occurring during the growth of metals and metal oxides were investigated, providing a much more precise understanding of how growth occurs. It is clear from the work that the deposition technique used plays a significant role on the resulting film quality and surface morphology and we are now able to provide an insight into the optimum conditions under which complete, crystalline layers can form