PhD, EngD and MSc by research theses (Cranfield Health)
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Browsing PhD, EngD and MSc by research theses (Cranfield Health) by Type "Thesis"
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Item Open Access Development of an affinity sensor for prostate cancer diagnosis(2010-11) Uludag, Yildiz; Tothill, Ibtisam E.Prostate carcinoma is a fatal malignancy and is a major cause of death in men in the population aged 55 and over. Early diagnosis of prostate cancer is very important for successful treatment of the disease. The increase in prostate specific antigen (PSA) levels in serum above the normal limits is the primary indication of prostate malignancy; therefore, PSA is used as a biomarker for the diagnosis and prognosis of prostate cancer. To date PSA testing still dominate as the best biomarker for prostate cancer detection even though it is not prostate specific. This project aims to develop a rapid, sensitive and specific biosensor to detect prostate cancer biomarkers in order to reduce the number of people who undergo invasive examinations for diagnosis. As part of the work, alternative surface chemistry techniques were investigated to optimise the sensors immobilisation capacity, such as the use of dendrimer monolayers and surface grafting polymers. It was found that dendrimer monolayers can be used to increase the sensor surface capacity by 1.5 times for surface plasmon resonance (SPR) sensor chips but not for quartz crystal microbalance (QCM) chips. A novel polymer (UVlink) surface was also fabricated and successfully used for biomolecule immobilisation for SPR and QCM chips. The QCMA-1 prototype biosensor was applied to develop PSA detection assays. Initially an immunoassay was constructed and optimised on the QCM sensor chip. A new buffer was then formulated to eliminate 98% of non-specific binding due to human sera proteins and this enabled the PSA detection assay to be performed in 75% human serum. Since QCMA-1 instrument is a prototype, to confirm the results the PSA assay was repeated using a commercial biosensor, Biacore 3000. This newly developed method showed a limit of detection of 0.29 ng mL"1 (in 75 % serum) with a linear dynamic detection range up to 150 ng mL"1 with Au nanoparticles employed for sensitivity enhancement using both instruments. With the achieved detection limit, it is possible to use the developed QCM and SPR assays for cancer detection in patient samples. PSA detection binding data fitted to 1:1 Langmuir binding model and KD calculated as 9.46 x IQ"10 M for the assay performed with Biacore 3000 instrument and using the QCMA-1 biosensor 5.56 x 10"10 M. These results were comparable and show that the affinity between PSA and PSA-capture antibody is in the region of 10'10 M and that QCMA-1 assay results are compatible with Biacore 3000 assay results. In the study incorporation of fPSA, cPSA and PSMA immunoassays to the prostate cancer detection test was also considered. However, the commercial available antibodies were formed to have low affinity for these markers. Patient samples collected from Bedford Hospital NHS Trust and control samples collected from Cranfleld University staff were tested and it was found that the developed QCMA-1 test clearly discriminates the PSA level of patients from the controls. A second biosensor platform based on label-free interdigitated capacitive sensing principle was also employed in the study. Interdigitated electrode (IDE) arrays were fabricated with conventional microelectronics-micromachining technologies on silicon wafers. Later IDE arrays were utilised for the detection of PSA using an LCR meter. This new prototype enabled capacitance reading of individual capacitors in turn and also enabled the detection to be performed in buffer solution. Antigen binding assays were developed to capture the PSA molecule in buffer solutions achieving a detection limit of 15.6 ng mL"1 with a linear dynamic detection range of 15.6 - 250 ng mL"1 PSAItem Open Access Ecological factors affecting growth and ochratoxin A production of Aspergillus section Nigri species on grapes(2006-05) Mitchell, D.; Magan, NareshThis study investigated the effects of interacting environmental factors on the ecophysiology of Aspergillus carbonarius and ochratoxin A (OTA) production, to facilitate the building of a database of knowledge for risk prevention. Spore germination and germ tube extension was faster on grape flesh than in vitro on grape- based media, or grape skin. Under optimum conditions (25-35°C; 0.90-0.99 water activity (aw)) 50% of conidia germinated in < 4.5 hours. Two dimensional profiles in relation to aw x temperature were constructed for rate of germination and germ tube extension for the first time. Mycelial growth was also influenced by aw, temperature, pH and modified atmospheres. Two dimensional temporal profiles of growth and OTA production were constructed based on information on strains from four different European countries. This showed that the optimum aw and temperature, and range were 0.98 aw, 30°C and 0.99-0.88 aw and 15-40°C, respectively. In contrast, for OTA production these were 0.95 aw, 15-20°C, and 0.99-0.93 aw and 15-35°C respectively. Environmental factors also influenced mycelial interactions between A. carbonarius and nine other common grape fungi. An Index of Dominance showed that A. carbonarius was very competitive against these species over a range of aw levels and temperatures. OTA production was generally suppressed by interactions at 0.95 aw with some inhibition of OTA production at 30°C at 0.99 and 0.95 aw. At 0.99 and 20°C all strains except two species produced a stimulation in OTA production. The efficacy of fungicides (5) and anti-oxidants (2) on growth and OTA production was limited at the concentration used. The only two fungicides that did suppress growth were Switch (cyprodinil and fludioxonil) and carbendazim. OTA production was stimulated at sub-optimal concentrations regardless of aw level used. Anti-oxidants were ineffective under all conditions tested. Modified atmospheres (0.03, 25 and 50% CO2) at different aw levels showed that there was very little long term effect on spore germination and germ tube extension, even at 50% CO2. Regardless of aW) OTA production was influenced more by the aw itself than CO2 concentrations. The efficacy of Sodium metabisulphite (NaMBS) at different aw levels (0.99, 0.97, 0.93 aw) on grape juice-based media at 25°C on germination and germ tube extension showed inhibition by >500 mg kg' 1 of NaMBS. Low concentrations (100, 250 mg kg'1) stimulated growth and OTA production. LD50 and LD90 ranges were determined for both growth and OTA production for the first time. Field sampling found that black aspergill! were present on grapes from setting to harvest with a significant increase in the population with grape ripening, and higher temperatures. The most commonly isolated OTA species from grapes was A. carbonarius with no Pénicillium verrucosum and only a few Aspergillus ochraceus isolated. In all the field sampling found no OTA present in any of the grape musts.Item Open Access Extraction of genetic network from microarray data using Bayesian framework(2007-04) Kumuthini, Judit; Bessant, Conrad; Setford, S.The aim of the work described in this thesis was to develop novel methods for the extraction of gene regulatory networks (GRN) from gene expression data, and use these methods to capture previously unknown relationships between genes in specific biological applications. This has been accomplished through the application of Bayesian Networks (BN) through minimum description length (MDL) and taboo search for parameter and structure learning respectively to three large scale microarray datasets from Saccharomyeces cerevisae, Escherichia coli and human stem cells. The application of BNs for modelling the well characterised yeast cell cycle demonstrated the efficacy of the techniques employed. Using the cDNA microarray data from the yeast cell cycle project by Spellman et a l (1998), this study succeeded in extracting many biologically plausible genetic relationships, which were supported by evidence from publicly available genome and literature databases. Two novel knowledge extraction techniques were applied; Target Node (TN) analysis and learning through simulation. Further, it was demonstrated how the addition of prior knowledge to the extracted network can improve the network structure extracted purely from experimental data. The second part of this thesis demonstrated how the BN approach could be adapted to a data set of very high dimensionality, specifically data from a 54,634 probe array used to monitor human adipose tissue. Genetic networks extracted included insulin receptor (IR) and Fatty acid binding proteins (FABP) families that play key roles in fatty acid uptake, transport, and metabolism In the final part of this thesis, the genome-wide GRNs of a prokaryotic expression system were extracted from novel oligo cDNA microarray data from E-coli K12 to identify metabolic stress responsive genes during recombinant protein production. Also, detailed analysis of known metabolic stress related genes and the genes that are directly or indirectly associated in the GRN were used to establish possible markers for host system exhaustion. In conclusion, the BN methods developed proved to be a powerful and effective means of extracting GRNs in a variety of applications.Item Open Access The hydraulic lung(2004-10) Prime, David; Turner, Anthony P. F.; Barr, H.A Hydraulic Lung has been designed and constructed. The Hydraulic Lung can inhale through a Dry Powder Inhaler (DPI) with a pre-determined level of inspiratory effort, and the characteristics of the inhalation profile generated, such as the peak pressure drop, peak flow rate and the flow acceleration are determined by the resistivity of the inhaler. The Hydraulic Lung has been used to explore the relationship between the level of inspiratory effort, the DPI resistivity and the resultant profile characteristics. A simple empirical equation has been found to describe the peak pressure drop achieved for any given level of inspiratory effort and device resistivity. This equation can be adapted to provide the equivalent peak inspiratory flow rate. A second simple empirical equation was found to describe the flow acceleration rate achieved under defined conditions of inspiratory effort and device resistivity. A clinical study has been performed to generate equivalent human inhalation data. A comparison between the relationships derived from the Hydraulic Lung data and the human inhalation data has demonstrated the validity of the key equation for pressure drop as a tool for predicHng human inhalation characteristics. The equation for flow acceleration rate was found to underestimate the flow accelerations achieved by human volunteers, but with slight modification could be used for this purpose. Correlations were established between the findings from this study and the work of earlier researchers in this area, which was based on clinical data alone. The Hydraulic Lung was also used as a practical tool for the evaluation of DPI performance in-vitro, including the behaviour of devices with variable resistivity which cannot easily be assessed using either standard pumps or sophisticated apparatus such as the Electronic Lung.Item Open Access An investigation into the effects of bacterial fermentation in autism(2006-11) Bilbao Montoya, Maria Pilar; Ritchie, Laurie; Hunter, J. O.Gastrointestinal symptoms have been observed to be present very often in autistic children. These symptoms are very similar to those observed in Irritable Bowel Syndrome (IBS). An investigation to investigate whether there is a similar cause for autism as well as IBS was done. Methods for the analysis of breath VOCs and urine organic acids were developed to be applied afterwards in the research of biomarkers of autism and IBS. Bacterial metabolites in urine and breath were identified by restricting the diets of healthy volunteers to each of two enteral feeds. The way enteral feeds work in the treatment of Crohn’s disease (CD) was investigated. Enteral feeds work by reducing bacterial activity in the gut, measurably reducing their metabolic products. The urine of IBS patients was analysed and bacterial compounds investigated. Some gastrointestinal symptoms are frequently misdiagnosed as IBS. According to the symptoms and treatment that is successful for these different groups of patients three subgroups can be made: “retention and overflow”, “musculoskeletal” and “abnormal fermentation” IBS. When the urine of these subgroups was observed, different biomarkers for the various subgroups could be identified. Patients with abnormal fermentation IBS have increased bacterial products in urine, suggesting an increase in bacterial activity in the colon. These biomarkers decreased after an exclusion or fibre-free diet and symptoms improved. In a further study, potential biomarkers for autism were identified. The origin of the majority of these compounds is unknown, although some of them are of known bacterial origin. A conclusion of this is that autistic children may exhibit abnormal fermentation since GI symptoms manifest at almost the same time as the autistic behavioural symptoms start and may be present from birth. Autistic children share many symptoms with IBS patients, although the biomarkers are different, showing that the GI symptoms observed in autistic children are not IBS in the children studied. More research needs to be done to determine the origin of the autism biomarkers discovered in urine.Item Open Access Nanofunctionalised orthopaedic implants(2018-01) Mirza, Farhan A.; Ge, YiDue to an aging population and younger patients presenting with musculoskeletal disorders, there is a need for orthopaedic implants with improved healing rates and longer implant life. Numerous research has developed implant surfaces with micro-topography and biomolecules to imitate the native extra cellular matrix (also known as biomimetic surfaces). This research has utilised such a biomimetic approach by immobilising the cell adhesive peptide, RGD (Arginine-Glycine-Aspartic Acid), to a titanium alloy Ti6Al4V surface. This research polymerised Hyperbranched Polyglycerol (HBPG) from the titanium surface using Ring Opening Multi-Branching Polymerisation (ROMBP). HBPG is a biologically compatible and non-toxic synthetic biopolymer, able to reduce non-specific protein adsorption, increase the titanium surface wetting (hydrophilicity), thereby limiting foreign body reactions. Extensive hydroxyl groups at the periphery of HBPG provides conjugation sites for biomolecule attachment. In this work the RGD peptide was conjugated to the polymer via a siloxane layer. This research developed a novel passivation solution for the preparation of the titanium alloy surface, using a mixture of hydrogen peroxide and nitric acid (a passivation mixture not used in the literature). This novel mixture was shown to etch the titanium surface, producing micro and nano surface features, both of which have been shown to improve cellular function in the literature. The hydrogen peroxide/nitric acid solution showed extensive oxidising ability on titanium, leading to the formation of reactable hydroxyl groups. Contact-angle measurements showed that the novel passivating solution produces a hydrophilic surface similar to that of peroxidation for 12-hours, but achieved in only 2-hours. In conjunction with the etching and oxidising abilities of hydrogen peroxide, the nitric acid reacts with the titanium surface, leading to the formation of a protective titanium oxide layer, enhancing corrosion resistance and improving biocompatibility. Biological investigations with the pre-osteoblast cell line MC3T3-E1 showed greater osteoblast cell attachment and adhesion strength, as well as improved bone matrix mineralisation on the passivated titanium surface functionalised with HBPG and the RGD peptide, compared to the raw and passivated titanium surfaces. Antibacterial testing of HBPG revealed substantially reduced bacterial cell colonies on the passivated/polymerised titanium surface, possibly arising from electrostatic and hydrophobic repulsion. This research has successfully developed a new titanium passivation solution (hydrogen peroxide/nitric acid) that can yield a contact-angle of around 35° in just 2-hours, rivalling the Piranha solution. The successful immobilisation of a cyclic RGD (cyclic-RGDfc) to a titanium surface functionalised with HBPG, has been shown in this research to drastically improve mineralised bone matrix production from the MC3T3-E1 cell line. This indicates earlier osseointegration of the implant may be possible, thereby improving patient healing times.Item Open Access A study of the cyber security awareness and use of protective cyber security practices in defence settings(2018-07) Sherchan, Sundar; Smy, VictoriaThe UK Strategic Defence and Security Review (2015), places ‘cyber’ in the highest category tier-one risk. The threats from cyberspace is ever increasing as UK Armed Forces is becoming increasingly dependent on its’ information systems and networks for daily business processes. Hardware and software technological defences are effective tools to protect our systems and networks, nonetheless these defences are useless if humans operators allows attackers to maliciously exploit our systems through use of social engineering techniques. There is currently no measurement framework in the R SIGNALS or the Army to assess basic cyber awareness and behaviour of soldiers and officers and benchmarking user cyber awareness maturity state. In this study, the author creates an innovative measurement framework that is utilised to measure cyber security awareness and behaviour in the R SIGNALS. The framework is an extension and adaptation of the government NCSC infographics for basic cyber security protective practices which in this study is split into five themes for measuring awareness (device safety, device backup, phishing, password and malware) and one theme for behaviour. The research adopts a quantitative positivist approach with using a questionnaire to measure human cyber awareness and behaviour. Study of human psychology models in the literature indicates that factors such as awareness and subsequent attitudes have direct influences on human behaviour. Results after codification and statistical analysis confirmed that technical trades in the R SIGNALS has better awareness of device safety, malware and phishing while cyber training was directly related to user behaviour and awareness of device safety. Overall user awareness in the R SIGNALS was found to be at Integrated level out of the five levels in the Community Cyber Security Maturity Model. The measurement framework is not limited to application to R SIGNALS and has the utility for other corps and organisations within the Army. Key future research recommendations included adding an attitude scale to the framework and having the correct sample to represent population variationItem Open Access Towards the development of an electrochemical immunosensor for the identification of transient ischemic attack via the labeless detection of biomedical markers.(2006) Barton, Andrew Charles; Higson, Seamus P. J.The thesis describes the development of generic platforms for labeless transduction of binding events. A particular application of the project is the labeless detection of biochemical markers for stroke, specifically transient ischemic attack, to facilitate rapid diagnosis and subsequent introduction of appropriate therapy within the required three-hour time frame. Novel biocomponents are under development for detection of these biochemical markers, and further demonstration of the generic nature of the technologies being developed will be realised via the use of a library of monoclonal antibodies as capture biocomponents. This work forms part of an investigation into labeless affinity immunosensor detection via impedimetric transduction utilising the technique of electrochemical impedance spectroscopy (EIS). Novel microelectrode arrays were fabricated by the sonochemical ablation of a homogeneous insulating layer of poly(o-phenylenediamine) exposing discrete areas of an underlying screen-printed conductive substrate. The receptor biocomponents are then subsequently coupled to the transducer by either electropolymerised entrapment or non-covalent affinity within and/or at the surface of conducting polymer ‘mushroom’ shaped protrusions for site-specific immobilisation. Upon exposure to a range of concentrations of antigen analyte solution, complex plane impedance analyses are used to relate the differing redox states of the polymer to possible charge transfer and / or related mechanisms between the immobilised antibody / antigen analyte and the polymer. Results show that signal differentiations were obtained, for a number of antibody / antigen species over a range of concentrations of clinical relevance (ng-pg/mf1), suggesting that the techniques employed offer a viable approach towards the labeless quantification of biochemical markers of transient ischemic attack (TIA).