Information processing applications driven by self-assembly and conformation dynamics of nucleic acids are possible. These underlying paradigms (self-assembly and conformation dynamics) are essential for natural information processors as illustrated by proteins. A key advantage in utilising nucleic acids as information processors is the availability of computational tools to support the design process. This provides us with a platform to develop an integrated environment in which an orchestration of molecular building blocks can be realised. Strict arbitrary control over the design of these computational nucleic acids is not feasible. The microphysical behaviour of these molecular materials must be taken into consideration during the design phase. This thesis investigated, to what extent the construction of molecular building blocks for a particular purpose is possible with the support of a software environment. In this work we developed a computational protocol that functions on a multi-molecular level, which enable us to directly incorporate the dynamic characteristics of nucleic acids molecules. To allow the implementation of this computational protocol, we developed a designer that able to solve the nucleic acids inverse prediction problem, not only in the multi-stable states level, but also include the interactions among molecules that occur in each meta-stable state. The realisation of our computational protocol are evaluated by generating computational nucleic acids units that resembles synthetic RNA devices that have been successfully implemented in the laboratory. Furthermore, we demonstrated the feasibility of the protocol to design various types of computational units. The accuracy and diversity of the generated candidates are significantly better than the best candidates produced by conventional designers. With the computational protocol, the design of nucleic acid information processor using a network of interconnecting nucleic acids is now feasible.

Ion channels are membrane proteins of interest for medical research and drug discovery, however a major bottleneck in obtaining functional measurements is the requirement to over-express the channel in-vivo. Cell-free (CF) protein expression is an alternative in-vitro approach capable of expressing proteins from a supplied DNA template - the method is fast, requires minimal apparatus and can be stabilised for the expression of membrane proteins by the addition of lipids or detergents. One drawback is the expense of commercial CF systems, however this can be economised by performing the reaction in microdroplets. This is attractive as microdroplets immersed in lipid-oil can be manipulated into contact to form a lipid bilayer, potentially allowing for ion channel expression and characterisation to be fully coupled. This study addresses the feasibility of achieving this goal by first investigating the stability of interdroplet bilayers formed in the presence of pre-incubated CF systems. Under these conditions the bilayers failed in <10 min, however a combination of diluting the mixture and adding vesicles was found to enable measurements of >30 min. The CF expression of the small prokaryotic potassium channel KcsA was then verified, in addition to the pore domain region of the eukaryotic hERG channel, where 20 ng/ul-74 ng/ul was expressed depending on the reaction conditions. Single-channel currents were subsequently obtained in interdroplet bilayers formed directly from the CF mixture, indicating that the channels were capable of self-inserting into the bilayer for measurements in both cases. The findings of this study support the feasibility of coupling the CF expression and electrical characterisation of ion channels in microdroplets and represent a progression toward the development of a high-throughput platform for screening novel pharmaceutical compounds.

Knowledge of the distribution and concentration of phytoplankton (microscopic algae that live in the oceans) is important, as this reflects and may modify environmentally important events such as pollution, climate change and carbon dioxide sequestration. Optical based flow cytometry has proved to be strategically important for the study of marine environments in general and of phytoplankton in particular because it enables rapid, simultaneous quantitative analysis of multiple optical properties of particles. Unfortunately, cost, complexity and size limit the widespread use of this technique for the study and monitoring of phytoplankton. The objective of this research was to solve some of the above mentioned problems by developing a small device for deployment on remote underwater vehicles (RUV) for in situ flow cytometric analysis of marine organisms. The core of the system was a lab-on-a-chip (LOC) device with a microfluidic channel with integrated optics and electrodes for the detection of the optical and electrical characteristics of phytoplankton. The work describes the design, fabrication, characterisation and testing of various LOC devices, together with data and analysis of results. The devices were able to clearly distinguish different populations of phytoplankton and other micro-particles on the basis of their optical and electrical properties. The systems had performance comparable with commercial instruments. Research also included bio-physical measurements (electro rotation and dielectrophoresis) of marine organisms to allow further modelling of the information obtained from the LOC cytometer.

The small size of integrated microfluidic systems offers many advantages such as cost, speed and portability for bioanalysis applications. However, due to the small scale the careful consideration of the transport of analytes in the bulk of the integrated microfluidic system and the interaction of analytes with surface immobilised analyte recognition molecules (receptors) is crucial for an efficient device operation. This thesis is concerned with the mathematical analysis and optimisation of the convective and diffusive transport of analytes and the analyte-receptor interaction in integrated microfluidic affinity systems with the aim of creating design guidelines for more efficient bioanalysis systems. The first part of this thesis considers device configurations where every analyte molecule can reach the surface immobilised receptors. In this case, which is important for sensing and separation applications, the transport-reaction model is solved analytically. This analytical solution is analysed for two bioanalytical applications: (i) affinity separation and (ii) affinity sensing. For fast analyte-receptor interactions, which are essential for affinity separation systems, the analysis of the analytical solution reveals simple expressions for the retention and the dispersion of the analyte due to the interaction with the receptors. With these expressions, which depend only on global device parameters, a framework for the design of multiplexed separation systems for the separation of proteins from complex sample mixtures is developed. Subsequently, the analytical solution of the transport-reaction model is used in the derivation of improved design strategies for microfluidic affinity sensors for the detection of analytes from small sample plugs. Three design strategies, which achieve a high capture fraction and a significantly increased uniformity of the bound analyte concentration over the sensor surface, are presented. The first two strategies rely on the variation of one device parameter, i.e. the flow velocity or the surface immobilised receptor concentration, as the analyte plug is flowed through the channel. The third approach is based on non-rectangular devices where the analyte plug is replenished by a narrowing of the flow channel. This third design strategy is applied to the redesign of a biosensor for the detection of low cytokine levels from small sample volumes. In the second part a novel microfluidic system for the generation of concentration gradients across microfluidic channels is developed. In this design the analytes are transported by surface groove induced secondary flow from the source to the sink stream. Numerical optimisations over the shape and size of the surface groove result in gradient generators which yield a well-defined linear or exponential concentration gradient across the width of the microfluidic channel. The resulting gradient generators have a much smaller footprint than conventional gradient generators and are thus more suitable for highly integrated lab-on-a-chip systems.

In recent years, the large-scale, untargeted studies of the compounds that serve as workers in the cell (proteins) and the small molecules involved in essential life-sustaining chemical processes (metabolites) have provided insights into a wide array of fields, such as medical diagnostics, drug discovery, personalised medicine and many others. Measurements in such studies are routinely performed using liquid chromatography mass spectrometry (LC-MS) instruments. From these measurements, we obtain a set of peaks having mass-to-charge, retention time (RT) and intensity values. Before further analysis is possible, the raw LC-MS data has to be processed in a data pre-preprocessing pipeline. In the alignment step of the pipeline, peaks from multiple LC-MS measurements have to be matched. In the identification step, the identity of unknown compounds in the sample that generate the observed peaks have to be assigned. Using tandem mass spectrometry, fragmentation peaks characteristic to a compound can be obtained and used to help establish the identity of the compound. Alignment and identification are challenging because the true identities of the entire set of compounds in the sample are unknown, and a single compound can produce many observed peaks, each with a potential drift in its retention time value. These observed peaks are not independent as they can be explained as being generated by the same compound. The aim of this thesis is to introduce methods to group these related peaks and to use these groupings to improve alignment and assist in identification during data pre-processing. Firstly, we introduce a generative model to group related peaks by their retention time. This information is used to influence direct-matching alignment, bringing related peak groups closer during matching. Investigations using benchmark datasets reveal that improved alignment performance is obtained from this approach. Next, we also consider mass information in the grouping process, resulting in PrecursorCluster, a model that performs the grouping of related peaks in metabolomics by their explainable mass relationships, RT and intensity values. Through a second-stage process that matches these related peak groups, peak alignment is produced. Experiments on benchmark datasets show that an improved alignment performance is obtained, while uncertainties in matched peaksets can also be extracted from the method. In the next section, we expand upon this two-stage method and introduce HDPAlign, a model that performs the clustering of related peaks within and across multiple LC-MS runs at once. This allows for matched peaksets and their respective uncertainties to be naturally extracted from the model. Finally, we look at fragmentation peaks used for identification and introduce MS2LDA, a topic model to group related fragmentation features. These groups of related fragmentation features potentially correspond to substructures shared by metabolites and can be used to assist data interpretation during identification. This final section corresponds to a work in progress and points to many interesting avenues for future research.

The purpose of this work was to study the effectiveness of diet derived antioxidants from wheat based aqueous cereal extracts under simulated gastrointestinal pH conditions and to monitor their effectiveness in modulating iron mediated oxidation which has been suggested as a risk factor in chronic disease. Wheat based breakfast cereals of composition ranging from whole grain to a refined flour product were analyzed for potential antioxidant effects. The breakfast cereals were extracted under aqueous conditions and the resulting extracts were tested for their ability to inhibit phosphatidylcholine liposome oxidation. The extent of oxidation was monitored by measuring the formation of thiobarbituric acid reaction substances (TBARS) and lipid peroxides. The aqueous extracts were analyzed using solvent extraction, molecular weight fractionation, phytate analysis, soluble fiber analysis, and total phenolics assay to determine the types of compounds responsible for the antioxidant activity. The state and modulation of iron before and after simulated gastrointestinal pH changes was monitored using atomic absorption spectrometry and the bathophenanthroline test. The aqueous extracts from the whole grain wheat and wheat bran breakfast cereals displayed considerable inhibition to lipid oxidation, while the wheat flour product was less effective. Following molecular weight fractionation the high molecular weight fraction was found to retain most of the antioxidative properties. The aqueous extracts subjected to solvent extraction with chloroform resulted in an organic extract containing non-polar compounds found in the aqueous extract. The antioxidant activity of this organic extract was minimal suggesting that the majority of compounds responsible for inhibiting oxidation are polar. Phytate analysis along with the use of a non-metal catalyst revealed that the antioxidant mechanism is not solely due to metal chelation. Precipitation and isolation of soluble fiber from the aqueous extracts were found to have no effect on oxidation. The total phenolics assay indicated that high concentrations of phenolics are present in the aqueous cereal extracts and appear to contribute to the inhibition of lipid oxidation. Simulated gastrointestinal pH conditions resulted in a significant increase in antioxidant activity for all aqueous cereal extracts including the low molecular weight (molecular mass <3,000 Da) fraction following ultrafiltration. Simulated gastrointestinal pH conditions resulted in the solubilization of iron in the cereals fortified with elemental iron. This increase in soluble iron was minimal and was found to exist complexed and not in a free ionic state. The solubilized iron following gastrointestinal pH conditions did not significantly effect the oxidation rate of phophatidy1choline liposomes in the model system. The chemical state of iron in the cereal product which was fortified with ferric phosphate was uneffected by the simulated gastrointestinal pH conditions and remained insoluble.

Sudden cardiac death kills 5-10 people per 10,000 population in Europe and the US each year. Individual propensity to arrhythmia and sudden cardiac death is typically assessed through clinical biomarkers. Variability in these biomarkers is a major challenge for risk stratification. Variability is observed at a wide range of spatio-temporal scales within the heart, from temporal fluctuations in ion channel behaviour, to inter-cell and inter-regional differences in ion channel expression, to structural differences between hearts. The extent to which variability manifests between spatial and temporal scales remains unclear but has a potentially crucial role in determining susceptibility to arrhythmia. In this dissertation we present a multi-scale study of the causes and consequences of variability in electrophysiology. At a sub-cellular level we demonstrate that, taking into account inter-individual variability in ion channel conductance, mRNA expression levels in failing human hearts predict the electrophysiological remodelling observed experimentally. On a tissue scale, we advocate the use of phenomenological models where information on subcellular processes is unavailable. We introduce a modification to a phenomenological model to capture beat-to-beat variability in action potential repolarisation recorded from four individual guinea pig myocytes. We demonstrate that, whilst temporal variability is dramatically reduced by inter-cell coupling, differences in their mean action potential duration may become apparent at a tissue level. The ventricular myocardium has a heterogeneous structure not captured by the simplified representation of conduction used above. In our final case study, we challenge a model of conduction by directly comparing simulations to optical mapping recordings of ventricular activation from failing and non-failing human hearts. We observe that good fits to experimental data are obtained only when endocardially bound structures are not in view, suggesting a role in conduction for these structures that are often ignored in cardiac simulations. Finally, we present future directions for the work presented. We make the case for reporting of inter-sample variability in experimental results and conclude that whilst variability may not always manifest across scales, its impact should be considered in both theoretical and experimental studies.

Electroencephalography (EEG) is the prevalent technique for monitoring brain function. It employs a set of electrodes on the scalp to measure the electrical activity of the brain. EEG is mainly used by researchers to study the brain’s responses to a specific stimulus - the event-related potentials (ERPs). Different types of unwanted signals, which are known as artefacts, usually mix with the EEG at any point during the recording process. As the amplitudes of the EEG and ERPs are very small (in the order of microvolts), they can be buried in the artefacts which have very high amplitudes in the order of millivolts. Therefore, contamination of EEG activity by the artefacts can degrade the quality of the EEG recording and may cause error in EEG/ERP signal interpretation. Several EEG artefact removal methods already exist in the literature and these previous studies have concentrated on manual or automatic detection of either one or, of a few types of EEG artefacts. Among the proposed methods, Independent Component Analysis (ICA) based techniques are commonly applied to successfully detect the artefacts. Different types of ICA algorithms have been developed, which aim to estimate the individual sources of a linearly mixed signal. However, the estimation criterion differs across various ICA algorithms, which may deliver different results.

Este estudo se fundamenta na construção de um modelo didático de formulação de perguntas que atenda às demandas educativas dos alunos nas aulas de Ciências e Biologia. Esse modelo didático tem como unidade central o discurso interativo entre professor e alunos para atribuição de significados compartilhados, que denominamos de discurso reflexivo. Consideramos que uma perspectiva de formação continuada reflexiva, investigativa e crítica possibilita que o professor implemente o modelo didático de formulação de perguntas provocando efeitos significativos nos processos cognitivos e interativos em sala de aula. Adotando o modelo didático de formulação de perguntas, os professores promovem modificações na sua postura pedagógica frente ao processo de ensino e aprendizagem das ciências, bem como re-significam as suas concepções sobre o currículo e seus elementos: conteúdos, objetivos educacionais, atividades de aprendizagem e avaliação. / This study is based upon the construction of a didactic model of questions formulation to meet the educational demands of students during the classes of Sciences and Biology. This didactic model has as its core unit the interactive discourse between the teacher and the students for the attribution of shared meanings, that we call reflexive discourse. We consider that the perspective of a reflexive, investigatory and critical continuing educational enables the teacher to implement the didactic model of questions formulation which will provoke meaningful effects in the cognitive and interactive process in the classroom. By adopting a didactic model of questions formulation, the teachers promote changes in their pedagogical attitude toward the teaching and learning sciences, and give new meaning to the their conceptions of the curriculum and its elements: contents, educational objectives, learning activities and evaluation.

O presente trabalho é a uma pesquisa empírica, de caráter qualitativo que tem como objeto de estudo a formação inicial de licenciandos de Ciências e Biologia. O objetivo central da investigação foi analisar se ela tem contribuído adequadamente para a tematização e construção de valores humanos e verificar sob a ótica dos licenciandos seu papel na formação ético-moral dos futuros estudantes e seu nível de preocupação sobre a dimensão ética dos saberes científicos e tecnológicos. O percurso metodológico consistiu em duas fases. Na primeira, licenciandos de Ciências e Biologia de três Instituições de Ensino Superior distintas responderam a um questionário referente à importância da formação ético-moral do estudante de ensino fundamental e médio e, para tanto, sua qualificação como docente. A segunda fase caracterizou-se pela aplicação de outro questionário com quatro casos com conteúdos dilemáticos e conflitos éticos nos quais os licenciandos identificaram ou opinaram sobre como os tratariam em sala de aula. Com base nas referências teóricas fornecidas pelo estudo na literatura voltada especificamente à educação ético-moral na perspectiva filosófica e psicológica, como também da Bioética de proteção, constatou-se que os licenciandos admitem que o aspecto ético-moral é fundamental para a formação do estudante, crêem que a escola, bem como outros ambientes sociais são co-participantes no desenvolvimento moral. Reconhecem a contribuição da disciplina de Ciências e Biologia como espaço de promoção de valores ético-morais e identificam assuntos que suscitam discussões éticas. Alguns obstáculos foram identificados que, direta e indiretamente, contribuem para o despreparo do professor em tratar dessas questões polêmicas, entre outros: a dificuldade de estimular e conduzir uma discussão, a insegurança quanto à perda do controle da classe, a não aceitação da divergência. Essas dificuldades podem ser atribuídas à sua trajetória de formação. Os resultados indicam a necessidade de implementação de novas estruturações para os cursos de formação destas disciplinas. A educação em Bioética favorece a inclusão da educação em valores no ensino de Ciências e Biologia, desde que os professores reconheçam que não podem se eximir de auxiliar seus futuros alunos a desenvolver habilidades necessárias para a reflexão sobre um problema e suas dimensões sociais, políticas e éticas requeridas na tomada de posição de todo o cidadão. / This is a qualitative empirical research, which aims at studying the initial training period of Science and Biology apprentice teachers. The main objective of the study was to analyze if this period has actually contributed properly to fostering human values, and, from the apprentice teachers point of view confirm his role in the ethical and moral training/education of his future students and his level of preoccupation with the ethical dimension of the scientific and technological knowledge. The methodological course of action consisted of two phases. In the first one, Science and Biology apprentice teachers from three different Universities answered a questionnaire about the importance of ethical and moral education for the Junior and High School student and consequently their qualification as teachers. The second phase consisted of another questionnaire containing four dilemmatic and conflicting cases in which the apprentice teachers identified or pointed out how the cases would be dealt with in class. Based on theoretical references provided by related literature concerning specifically the ethical and moral education from the philosophical and psychological perspectives, as well as the Bioethics of protection, the apprentice teachers admit that ethical and moral aspects are essential for the students education. Also, teachers believe that the schools, as well as other social environments are co-participants in the moral development of their students. Teachers also identify issues that raise ethical discussions and recognize the contribution of Sciences and Biology as a way of promoting ethical and moral values. Some obstacles were detected, which directly or indirectly contribute to the teachers lack of preparation regarding those polemic issues, such as: the difficulty in stimulating and guiding a discussion, the lack o self-assurance in handling the group, the non-acceptance of divergencies. Such difficulties may stem from their educational background. The results show the need to implement new structures for the training course of these subjects. Bioethics education favors the inclusion of values education when teaching Science and Biology, provided that teachers acknowledge they cannot do without helping students develop the necessary skills to reflect on a problem in its social, political and ethic dimensions. These skills are essential for every citizen when taking a stand.