Development of novel antibody-based diagnostics for the early and rapid detection of cardiac markers
Year: 2010
Type: dissertation
Abstract: The principal objective of this research was the production, characterisation and application\nof antibodies for the detection of both C-reactive protein (CRP) and Myeloperoxidase\n(MPO), two promising candidates in the field of cardiac diagnostics. CRP is a highly stable\nacute phase reactant, produced in the liver in response to interleukin (IL)-6. In January 2003\nboth the Centres for Disease Control and Prevention (CDC) and the American Heart\nAssociation (AHA) recommended CRP as the inflammatory marker of choice for assessment\nof cardiovascular risk. The other biomarker under investigation was MPO, a mediator\nenzyme secreted by inflammatory cells such as activated neutrophils and monocytes. High\nlevels of MPO have been reported in atherosclerotic plaques and accumulating evidence\nindicates that MPO may have a causative role in plaque vulnerability. It has been suggested\nthat these markers may even be complementary in the assessment of patient cardiac risk. As a\nmarker of disease activity and vascular inflammation, CRP detection is associated with longterm risk stratification while MPO, being a marker of plaque instability and neutrophil\nactivation, may be prove to be a key marker for short term stratification.\nA key feature of this work was to ensure the isolated antibodies had a sufficient level of both\nfunctional affinity and specificity for the development of novel diagnostic formats and assay\nplatforms. Both classic polyclonal antibody generation and ‘state-of-the-art’ phage display\ntechnology were employed for the generation of CRP- and MPO-specific antibodies and\nrecombinant fragments. The immune systems of mice, rabbits and chickens were investigated\nin this work, which allowed for the identification of the main advantages and disadvantages\nassociated with each of the host immune systems and their capacity for generating specific\nand sensitive antibodies. It was observed that the avian immune system was found to be the\nmost effective system for the generation of functional and soluble antibodies.\nThe CRP-specific antibodies were incorporated into three novel biosensor platforms for their\nfinal characterisation. In addition, a novel lateral flow immunoassay (LFIA) for the rapid\ndetection of MPO was developed, using a recombinant antibody isolated from a MPOimmune library. Spiked samples of depleted human serum were employed in both the\nbiosensor and LFIA systems, thus validating the detection of the analytes in a realistic sample\nmatrix
Author: Barry J. McDonnell
Language: English
Subfield: Immunology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2395064507