Chronic obstructive pulmonary disease (COPD) encompasses the diseases of chronic bronchitis and emphysema, both of which can cause the main feature of this disease; airflow obstruction. COPD is the third leading cause of mortality in the world, and although cigarette smoking is the main risk factor of the disease, other environmental factors and genetic disorders can also play a role. The pathogenesis of COPD is currently still poorly documented, which results in poor diagnosis and treatment for the disease. The involvement of autoimmunity in the pathogenesis of the COPD is becoming more apparent; in recent studies, high levels of circulating autoantibodies have been detected in patients with COPD, suggesting that this lung disorder may have an autoimmune component. Therefore, research has focused on identifying a set of autoantigens known to be associated with other autoimmune disorders that may play a role in the pathogenesis of this disease. The aim of this study was to develop antigen microarrays for profiling antigen specific-autoantibodies in the serum of patients with COPD, as a diagnostic tool at the early stage of the disease. To achieve the project aim, we have employed antigen microarrays. Before applying this technique, quality control experiments were performed to optimize the procedure. This included testing different slide surface coatings, investigating numerous blocking buffers and methods to amplify the signal intensities, as well as a series of validatory tests to determine if the assay was both accurate and reproducible. The optimization results showed that a combination of amino-silaine coated slides with I Block buffer and Genisphere amplifier performed well due to the high signal detection and low background observed. Validation for optimized technique was achieved and showed no cross-reactivity occurred, an acceptable limit of the coefficient of variation (CV %), and a significant correlation between ELISA and the antigen microarray platform, with microarray giving a bigger dynamic range of signals. The results for the limit of detection (LOD) were applied and the cut off for a positive autoantibody response was taken from the 95th percentile of the healthy non-smokers control group. The results for the investigation of the different 39 autoantigens showed that there was a significant increase in reactivity for both the IgG and IgM autoantibodies in the COPD group compared to the control for CENP-B, collagen5, RNP/sm, La ssb, histone, ro-52 and SCL-70, which suggests these antigens have the potential to be used as diagnostic biomarkers for the detection of COPD. These results also showed that a number of healthy smokers produced an increased reactivity to the same autoantigens as the COPD patients, which suggests there is potential susceptibly of these smokers to develop COPD. In conclusion, this thesis developed a rapid, inexpensive, broad-spectrum antigen microarray technology, which could have a pivotal future role in the early diagnosis of COPD.

Neutralising antibodies are an essential component in the immune response to virus infection and are often effective at preventing or attenuating viral diseases. The administration of cocktails of potently neutralising monoclonal antibodies (nAb) to abrogate viral infections has been demonstrated as an effective treatment to control disease progression caused by a diverse range of viruses. The mechanism of neutralisation is mediated predominantly by nAb binding to glycosylated envelope proteins displayed on the surface of virus particles, thereby preventing binding to host cell membrane bound receptor proteins and neutralising the entry process. Although a robust nAb response is elicited following infection with a number of viruses, many human viruses have evolved mechanisms to effectively evade the humoral immune response. The emergence or re-emergence of pathogenic human viruses poses a significant threat to the global population. The Ebolavirus epidemic of 2014-2015 arose from the emergence of a known virus in a new area, which rapidly spread through a naïve population with devastating consequences, resulting in 11,323 fatalities from a total of 28,646 confirmed cases of infection. A rapid induction of potent, broadly nAbs was a possible pathway for infection control in the 17,323 survivors of the epidemic. The isolation of potent nAbs from seroconverted individuals following antigenic exposure provides the potential to develop novel therapeutics for administration within a region affected by an epidemic. To this end, the aim of this project was to develop a platform of methods to isolate human monoclonal antibodies directly from peripheral blood through the isolation of memory B-lymphocytes and subsequent stimulation to secrete antibodies using an in vitro culture system. Identification of DNA sequences encoding antibody heavy and light chain variable domains (VH and VL respectively), allowed for the production of recombinant antibody following the sub-cloning of VH and VL regions into plasmid expression vectors to provide a proof of principle of platform function. Secondly, the introduction of functional screens to the platform would provide a method to identify antibodies derived from memory B-cells with the ability to neutralise Ebolavirus pseudo-particles using in vitro entry inhibition assays.