Simple and Rapid On-site molecular detection of Mycobacterium tuberculosis

描述（由申请人提供）：SBIR第一阶段申请将集中于开发一种简单廉价的手持式电池供电的生物传感器，该生物传感器集成了微流体和分子生物学，可快速检测结核分枝杆菌（人类结核病（TB）的病原体）。由于每年有多达900万新发结核病病例和约200万例死亡，以及资源有限的国家在准确诊断结核病方面遇到的困难，世界卫生组织敦促开发新的诊断平台以检测M。结核资源有限的国家依赖于100年前缺乏灵敏度的显微镜方法，是成功产品的主要市场。此外，通过航空旅行将耐多药结核病传播到工业化国家的风险使其成为全球健康问题。由于每年花费近10亿美元进行结核病检测，因此易于使用的结核分枝杆菌检测的商业市场潜力很大。结核病是迫在眉睫的，并将对改善全球健康问题产生广泛影响。 为了利用现代分子生物学检测方法，而不需要昂贵的设备和耗时的分析步骤，我们建议利用两组不同的寡核苷酸探针互补的M。结核病rRNA序列。一组将与磁珠结合，而第二组将与包封酶底物的脂质体缀合。所得杂交复合物将通过磁体固定在微流体装置内。然后将脂质体裂解，允许现在释放的酶底物驱动电化学反应，该电化学反应将通过集成的叉指型超微电极阵列（IDUA）进行定量。因此，IDUA将生物信号转换为数字电子信号。这些目标和目的的实现将允许直接检测M。结核病rRNA，而不需要耗时的富集或基因扩增步骤。此外，预计这种易于使用的诊断测试将允许由不需要广泛培训和分子生物学背景的个人进行近患者测试。 成功完成拟议的第一阶段工作将导致一个独特的生物传感器，能够快速检测结核分枝杆菌，结核病的病原体。由于世界上超过三分之二的结核病例是在缺乏财政或技术资源来准确诊断新的结核病例或监测现有病例的治疗的资源有限的国家发现的，这种生物传感器将对改善生活在这些国家的个人的健康产生直接和巨大的影响。此外，由于TB可通过空气传播有效地传播，因此在世界上具有高感染率的那些地区控制TB的能力也将降低TB传播的风险，包括由M的多药耐药菌株引起的风险。结核病，工业化国家。

DESCRIPTION (provided by applicant): This SBIR Phase I application will focus on the development of a simple and inexpensive, handheld, battery-operated biosensor integrating microfluidics and molecular biology to rapidly detect Mycobacterium tuberculosis, the etiologic agent of tuberculosis (TB) in humans. With as many as 9 million new cases of TB and approximately 2 million deaths per year and the difficulties encountered in resource-limited countries to accurately diagnose TB, the World Health Organization has urged the development of new diagnostic platforms for the detection of M. tuberculosis. Relying upon 100-year old microscopic methods that lack sensitivity, resource-limited countries represent a major market for a successful product. Furthermore, the risk of transmission of multi-drug resistant TB to industrialized nations via the availability of air travel makes this a global health issue. With almost $ 1.0 Billion spent every year to perform TB testing, the commercial market potential for an easy-to-use test for M. tuberculosis is immediate and will have broad implications for improving a worldwide health issue. In order to exploit modern molecular biology detection methods without imposing a need for expensive equipment and time-consuming analytical steps, we propose to utilize two different sets of oligonucleotide probes complementary to a M. tuberculosis rRNA sequence. One set will be bound to magnetic beads while a second set will be conjugated to liposomes that encapsulate enzyme substrates. The resulting hybridization complex will be immobilized via a magnet within a microfluidic device. Liposomes will then be lysed allowing the now released enzyme substrates to drive an electrochemical reaction that will be quantitated by an integrated interdigitated ultramicroelectrode array (IDUA). Thus, the IDUA will transduce the biological signal into a digital electronic signal. The accomplishment of these goals and objectives will permit the direct detection of M. tuberculosis rRNA without the need for time-consuming enrichment or gene amplification steps. Furthermore, it is anticipated that this easy-to-use diagnostic test will permit near-patient testing by individuals who will not require extensive training and background in molecular biology. Successful completion of the proposed Phase I effort will result in a unique biosensor, capable of rapidly detecting Mycobacterium tuberculosis, the causative agent of TB. Inasmuch as more than two-thirds of the world's TB cases are found in resource-limited countries that lack the financial or technical resources to accurately diagnose new TB cases or monitor treatment of existing cases, such a biosensor will have an immediate and dramatic impact on improving the health of individuals living in such countries. Furthermore, since TB can be efficiently spread by airborne transmission, the ability to control TB in those portions of the world with high infection rates will also reduce the risk of spreading TB, including that caused by multi-drug resistant strains of M. tuberculosis, to industrialized nations.