Uropathogen Detection Using DNA Biosensors

描述（由申请人提供）：尿路感染是最常见的 常见的泌尿系统疾病在美国，是一个主要的原因， 发病率和保健支出。这个生物工程研究伙伴关系 建议涉及开发一个微机电系统， 基因型检测和种属特异性鉴定 一个时间框架（从样品收集到读出的5-10分钟）， 即时诊断和治疗。该提案的重点是生产 一个独立的微生物病原体检测设备，并检查其 使用临床尿液样本的性能。加州大学洛杉矶分校的研究提供了两个关键 这些技术进步使得泌尿病原体传感器的开发成为可能。 第一个是用于样品处理的微流体技术。第二个是 电化学微传感器，其允许超灵敏地检测特定的 DNA-RNA或DNA-DNA杂交事件，无需靶点 放大这个项目已经开发了一年多， 多学科的努力，包括微流体领域的领导者， 微传感器技术、分子微生物学、儿科学和生物数学。 具体目标1描述了微流体研究将如何应用于 用于泌尿系病原体浓缩、微量混合的错流过滤器的开发 用于处理泌尿病原体核酸和清洗传感器表面。 具体目标2涉及微传感器阵列的制造， 链霉亲和素自组装单层，以及寡核苷酸探针的测试 用于微传感器上尿路病原体rRNA和mRNA的电化学检测 面具体目标3将涉及微流体和 传感器组件和测试其分析有效性的模拟和实际 尿液样本具体目标4将涉及器械的批量制造 以及尿传感器结果与 尿路感染的临床相关因素。

DESCRIPTION (provided by the applicant): Urinary tract infection is the most common urological disease in the United States and is a major cause of patient morbidity and health-care expenditure. This Bioengineering Research Partnership proposal involves development of a microelectromechanical system for the genotypic detection and species-specific identification of uropathogens within a time frame (5-10 minutes from sample collection to readout) that would enable point-of-care diagnosis and treatment. The focus of this proposal is to produce a self-contained microbial pathogen detection device and to examine its performance using clinical urine samples. Research at UCLA has provided two key technological advances that make development of a uropathogen sensor feasible. The first is microfluidics for sample processing. The second is an electrochemical microsensor which allows ultrasensitive detection of specific DNA-RNA or DNA-DNA hybridization events, without the need for target amplification. This project has been in development for over a year involving a multidisciplinary effort including leaders in the fields of microfluidics and microsensor technology, molecular microbiology, pediatrics and biomathematics. Specific Aim 1 describes how microfluidics studies will be applied to development of a crossflow filter for uropathogen concentration, micromixing for processing of uropathogen nucleic acids, and washing of the sensor surface. Specific Aim 2 involves fabrication of the microsensor array, development of a streptavidin self-assembled monolayer, and testing of oligonucleotide probes for electrochemical detection of uropathogen rRNA and mRNA on the microsensor surface. Specific Aim 3 will involve integration of the microfluidics and sensor components and testing of its analytic validity on simulated and actual urine specimens. Specific Aim 4 will involve batch fabrication of the device and clinical examination of the association between urosensor results and clinical correlates of urinary tract infection.