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Title page for ETD etd-03222018-182535
|Type of Document
||Markwalter, Christine Franzel
||Strategies for Improving Diagnosis of Malaria and Schistosomiasis in Low-Resource Settings
|David W. Wright
|David E. Cliffel
|James E. Crowe, Jr.
|Janet E. Macdonald
- Histidine-rich protein 2
- Plasmodium lactate dehydrogenase
|Date of Defense
Infectious diseases claim tens of millions of lives globally each year and greatly impact the quality of life of many others. Low- and middle-income countries bear the vast majority of the global burden of infectious disease morbidity and mortality. Development and implementation of accurate and appropriate diagnostic tools are critical for case management, surveillance, and elimination campaigns. However, in settings lacking advanced healthcare infrastructure, diagnosis of infectious diseases presents unique challenges. In this work, each component of the conventional diagnostic format (sample matrix, molecular recognition, and signal generation) was examined and optimized with the goal of improving diagnosis of malaria and schistosomiasis in endemic settings. First, sample matrix effects were evaluated and mitigated. For malaria, the extent to which the host immune response affects detection of malarial biomarker histidine-rich protein 2 (HRP2) in a low-transmission setting was determined, and strategies for reducing this potential matrix effect were evaluated. For schistosomiasis, novel dendrimer-coated magnetic particles were developed to capture, purify, and concentrate the circulating anodic antigen (CAA) biomarker from its variable urine matrix before application to an ultrasensitive lateral flow assay. Next, the affinities of monoclonal antibodies for two malarial biomarkers, HRP2 and Plasmodium lactate dehydrogenase (pLDH), were screened using biolayer interferometry. It was found that kinetic parameters of individual antibodies against HRP2, but not pLDH, predicted performance of antibody pairs in a sandwich enzyme-linked immunosorbent assay (ELISA) format. These data informed the development of highly sensitive magnetic bead-based assays for both biomarkers, ultimately resulting in a rapid multiplexed assay in which pLDH and HRP2 detection could be completed in less than 1 hour with detection limits an order of magnitude better than commercially available ELSIA kits. Finally, the assay was applied to the characterization of pLDH and HRP2 clearance patterns in patients living in a highly endemic region of Zambia, demonstrating the sensitivity and clinical utility of the developed assay.
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