Electrochemiluminescence: insights into the mechanisms and nanomaterial-based applications

Biomarkers are biological indicators of human health conditions. Their ultra-sensitive quantification is of paramount importance in clinical monitoring and early disease diagnosis. Biosensors are simple and easy-to-use analytical devices and, in their world, electrochemiluminescence (ECL) is one of the most promising analytical techniques that needs an ever-increasing sensitivity for improving its clinical effectiveness. Scope of this project was the investigation of the ECL generation mechanisms for enhancing the ECL intensity also through the identification of suitable nanostructures. The combination of nanotechnologies, microscopy and ECL has proved to be a very successful strategy to improve the analytical efficiency of ECL in one of its most promising bioanalytical approaches, the bead-based immunoassay. Nanosystems, such as [Ru(bpy)3]2+-dye-doped nanoparticles (DDSNPs) and Bodipy Carbon Nanodots, have been used to improve the sensitivity of ECL techniques thanks to their advantageous and tuneable properties, reaching a signal increase of 750% in DDSNPs-bead-based immunoassay system. In this thesis, an investigation of size and distance effects on the ECL mechanisms was carried out through the innovative combination of ECL microscopy and electrochemical mapping of radicals. It allowed the discovery of an unexpected and highly efficient mechanistic path for ECL generation at small distances from the electrode surface. It was exploited and enhanced through the addition of a branched amine DPIBA to the usual coreactant TPrA solution for enhancing the ECL efficiency until a maximum of 128%. Finally, a beads-based immunoassay and an immunosensor specific for cardiac Troponin I were built exploiting previous results and carbon nanotubes features. They created a conductive layer around beads enhancing the signal by 70% and activating an ECL mechanism unobserved before in such systems. In conclusion, the combination of ECL microscopy and nanotechnology and the deep understanding of the mechanisms responsible for the ECL emission led to a great enhancement in the signal.

Architectures for Context Aware Services in Smart Environments

This thesis deals with Context Aware Services, Smart Environments, Context Management and solutions for Devices and Service Interoperability. Multi-vendor devices offer an increasing number of services and end-user applications that base their value on the ability to exploit the information originating from the surrounding environment by means of an increasing number of embedded sensors, e.g. GPS, compass, RFID readers, cameras and so on. However, usually such devices are not able to exchange information because of the lack of a shared data storage and common information exchange methods. A large number of standards and domain specific building blocks are available and are heavily used in today's products. However, the use of these solutions based on ready-to-use modules is not without problems. The integration and cooperation of different kinds of modules can be daunting because of growing complexity and dependency. In this scenarios it might be interesting to have an infrastructure that makes the coexistence of multi-vendor devices easy, while enabling low cost development and smooth access to services. This sort of technologies glue should reduce both software and hardware integration costs by removing the trouble of interoperability. The result should also lead to faster and simplified design, development and, deployment of cross-domain applications. This thesis is mainly focused on SW architectures supporting context aware service providers especially on the following subjects: - user preferences service adaptation - context management - content management - information interoperability - multivendor device interoperability - communication and connectivity interoperability Experimental activities were carried out in several domains including Cultural Heritage, indoor and personal smart spaces – all of which are considered significant test-beds in Context Aware Computing. The work evolved within european and national projects: on the europen side, I carried out my research activity within EPOCH, the FP6 Network of Excellence on “Processing Open Cultural Heritage” and within SOFIA, a project of the ARTEMIS JU on embedded systems. I worked in cooperation with several international establishments, including the University of Kent, VTT (the Technical Reserarch Center of Finland) and Eurotech. On the national side I contributed to a one-to-one research contract between ARCES and Telecom Italia. The first part of the thesis is focused on problem statement and related work and addresses interoperability issues and related architecture components. The second part is focused on specific architectures and frameworks: - MobiComp: a context management framework that I used in cultural heritage applications - CAB: a context, preference and profile based application broker which I designed within EPOCH Network of Excellence - M3: "Semantic Web based" information sharing infrastructure for smart spaces designed by Nokia within the European project SOFIA - NoTa: a service and transport independent connectivity framework - OSGi: the well known Java based service support framework The final section is dedicated to the middleware, the tools and, the SW agents developed during my Doctorate time to support context-aware services in smart environments.

Semantic coordination through programmable Tuple spaces

Two of the main features of today complex software systems like pervasive computing systems and Internet-based applications are distribution and openness. Distribution revolves around three orthogonal dimensions: (i) distribution of control|systems are characterised by several independent computational entities and devices, each representing an autonomous and proactive locus of control; (ii) spatial distribution|entities and devices are physically distributed and connected in a global (such as the Internet) or local network; and (iii) temporal distribution|interacting system components come and go over time, and are not required to be available for interaction at the same time. Openness deals with the heterogeneity and dynamism of system components: complex computational systems are open to the integration of diverse components, heterogeneous in terms of architecture and technology, and are dynamic since they allow components to be updated, added, or removed while the system is running. The engineering of open and distributed computational systems mandates for the adoption of a software infrastructure whose underlying model and technology could provide the required level of uncoupling among system components. This is the main motivation behind current research trends in the area of coordination middleware to exploit tuple-based coordination models in the engineering of complex software systems, since they intrinsically provide coordinated components with communication uncoupling and further details in the references therein. An additional daunting challenge for tuple-based models comes from knowledge-intensive application scenarios, namely, scenarios where most of the activities are based on knowledge in some form|and where knowledge becomes the prominent means by which systems get coordinated. Handling knowledge in tuple-based systems induces problems in terms of syntax - e.g., two tuples containing the same data may not match due to differences in the tuple structure - and (mostly) of semantics|e.g., two tuples representing the same information may not match based on a dierent syntax adopted. Till now, the problem has been faced by exploiting tuple-based coordination within a middleware for knowledge intensive environments: e.g., experiments with tuple-based coordination within a Semantic Web middleware (surveys analogous approaches). However, they appear to be designed to tackle the design of coordination for specic application contexts like Semantic Web and Semantic Web Services, and they result in a rather involved extension of the tuple space model. The main goal of this thesis was to conceive a more general approach to semantic coordination. In particular, it was developed the model and technology of semantic tuple centres. It is adopted the tuple centre model as main coordination abstraction to manage system interactions. A tuple centre can be seen as a programmable tuple space, i.e. an extension of a Linda tuple space, where the behaviour of the tuple space can be programmed so as to react to interaction events. By encapsulating coordination laws within coordination media, tuple centres promote coordination uncoupling among coordinated components. Then, the tuple centre model was semantically enriched: a main design choice in this work was to try not to completely redesign the existing syntactic tuple space model, but rather provide a smooth extension that { although supporting semantic reasoning { keep the simplicity of tuple and tuple matching as easier as possible. By encapsulating the semantic representation of the domain of discourse within coordination media, semantic tuple centres promote semantic uncoupling among coordinated components. The main contributions of the thesis are: (i) the design of the semantic tuple centre model; (ii) the implementation and evaluation of the model based on an existent coordination infrastructure; (iii) a view of the application scenarios in which semantic tuple centres seem to be suitable as coordination media.

A clustering method for robust and reliable large scale functional and structural protein sequence annotation

Bioinformatics, in the last few decades, has played a fundamental role to give sense to the huge amount of data produced. Obtained the complete sequence of a genome, the major problem of knowing as much as possible of its coding regions, is crucial. Protein sequence annotation is challenging and, due to the size of the problem, only computational approaches can provide a feasible solution. As it has been recently pointed out by the Critical Assessment of Function Annotations (CAFA), most accurate methods are those based on the transfer-by-homology approach and the most incisive contribution is given by cross-genome comparisons. In the present thesis it is described a non-hierarchical sequence clustering method for protein automatic large-scale annotation, called “The Bologna Annotation Resource Plus” (BAR+). The method is based on an all-against-all alignment of more than 13 millions protein sequences characterized by a very stringent metric. BAR+ can safely transfer functional features (Gene Ontology and Pfam terms) inside clusters by means of a statistical validation, even in the case of multi-domain proteins. Within BAR+ clusters it is also possible to transfer the three dimensional structure (when a template is available). This is possible by the way of cluster-specific HMM profiles that can be used to calculate reliable template-to-target alignments even in the case of distantly related proteins (sequence identity < 30%). Other BAR+ based applications have been developed during my doctorate including the prediction of Magnesium binding sites in human proteins, the ABC transporters superfamily classification and the functional prediction (GO terms) of the CAFA targets. Remarkably, in the CAFA assessment, BAR+ placed among the ten most accurate methods. At present, as a web server for the functional and structural protein sequence annotation, BAR+ is freely available at http://bar.biocomp.unibo.it/bar2.0.