Chemical and Biomolecular Engineering - Theses
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ItemApplication of Advanced Fluorescence Imaging Techniques for Intracellular Tracking of Nano-biomaterials( 2021)The engineering and intracellular delivery of nanoparticles with tailored structural, functional and therapeutic properties is challenging due to the interactions of nanomaterials with complex and dynamic biological systems. Additionally, the clinical translation of nanoparticle-based chemotherapeutics is hampered by the poor capacity of 2D cell monolayer culture to mimic in vivo tumour microenvironment and cell-cell interactions. To overcome these biological barriers and enable the clinical translation of nanoparticles, a thorough investigation of nanoparticle-cell interactions in complex biological environments is of paramount importance. For this purpose, novel advanced fluorescence techniques enable the study of nanoparticles structure and functional properties inside the cells with improved spatial and temporal resolution. Additionally, the development of complex 3D cell culture systems mimicking tumour tissue could provide a novel method to predict the in vivo behaviour of nanoparticle-based chemotherapeutics and cellular response to the treatment. Herein DNA- and sugar-based nanoparticles have been developed as platforms for the detection of molecular targets and delivery of drugs within cells and in complex biological settings. Specifically, fluorescence resonance energy transfer microscopy, fluorescence correlation spectroscopy, fluorescence lifetime imaging microscopy and multicolour single-molecule localization microscopy were employed to probe the specific binding of the DNA nanosensor to the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) and the nanocomplexation of glycogen with albumin. The intracellular trafficking and the activity of drug-loaded nanoparticles were investigated in 2D and 3D static and dynamic cell culture systems. The biological activity of glycogen-albumin nanoparticles was investigated in a 3D tumour microtissue obtained by co-culturing BT474, NIH-3T3 and RAW264.7 cells in a U-Cup perfusion bioreactor device. The interactions of glycogen-albumin nanoparticles with peripheral blood mononuclear cells isolated from human blood and nanoparticles in vivo biodistribution in mice were also analyzed. This study aims to gain an understanding of the bio-nano interactions in various biological systems and highlights the importance of combining multiple fluorescence techniques and complex models for monitoring the intracellular behaviour of nanomaterials and accurately predicting their in vivo behaviour.
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ItemEngineering catalytic organic-inorganic materials for sensing applications( 2020)Nanostructured hybrid organic-inorganic materials are a unique class of materials showing distinctive properties that have attracted high interest due to their diverse applications in the fields of energy, environment and medicine. In particular, hybrid materials are promising candidates for sensing applications due to the tunable chemical, structural and functional properties of the organic and inorganic components. Hence, the engineering of novel nanostructured catalytic organic/inorganic materials provides opportunities for the fabrication of advanced nanodevices for biosensing. In this thesis, novel hybrid materials have been prepared and their electrocatalytic, catalytic, and optical properties explored. First, nanostructured electrocatalytic microparticles were synthesized in mild conditions and used with an organic binding agent to prepare carbon electrodes applied in the detection of glucose in biologically relevant media. Second, hierarchically structured hybrid particles displaying enzyme-like catalytic activities were synthesized and used to prepare high-throughput micro-reactors for the detection of bioanalytes via a hybrid organic-inorganic cascade reaction. Finally, a natural occurring polysaccharidic nanoparticle, i.e. glycogen, was engineered to impart adhesive functional properties to a hybrid film and used for the coating of various substrates with different chemical composition. These hybrid coatings embedding metal nanoparticles were employed as catalytic and optically active functional interfaces.