Development of diagnostic system using molecular technology

Abstract

In recent years, many approaches have been developed to detect biomolecules for diagnosis for various diseases. The success of these approaches largely relies on the integration of interdisciplinary knowledge of molecular biology, chemistry and physics. The detection system in molecular diagnostics demands high sensitivity and specificity by employing various materials and platforms such as microarray-based techniques and nanomaterials-based biosensor. Additionally, as all diagnostic devices and products are trending toward high throughput screening and miniaturization, electrical approaches as well as optic approaches come into the spotlight and its studies are vigorously progressed worldwide. This thesis introduces the development of various diagnostic tools using molecular technology for clinical disease. The value and practicality of these diagnostic systems developed in this thesis is enhanced by applying the clinical samples. First, DNA microarray for microbial pathogen detection was developed by considering high prevalence rate and/or difficulty of cultivation of pathogen. I directly determined the sequences of 23S ribosomal DNA (rDNA) and 16S-23S rDNA intergenic spacer region (ISR) for 19 bacteria with the limited or no availability of their sequences in public databases,facilitating the advance of various assays for pathogen detection. Employing bacteria-universal probes as well as species-specific probes in DNA microarray system enables to reduce false negative results by detecting all bacteria. As a result of the validation using 515 clinical isolates, the DNA microarray allowed efficient detection of bacterial pathogens with the specificities of 100% with sensitivities of 100% as well except for two pathogens. Furthermore, the detection of pathogen using DNA microarray was expanded up to 52 bloodborne pathogen, so far the largest number of pathogens. The microarrays which were applied to 112 blood culture positive samples, showed also successful i...