Integrated molecular diagnostic system for the point-of-care

描述（由申请人提供）：该STTR-AT-NIAID旨在基于波士顿大学Catherine Klapperich实验室的研究开发一种集成核酸系统。BU芯片实验室包括一个微型固相萃取（<SPE）柱、瓣阀和疏水排气口，以控制流体通过微通道的运动，以及用于实验重复和控制反应的多个反应室。Klapperich博士的实验室已经成功地使用BioHessel的等温扩增过程在芯片中进行了核酸扩增。Biohelix的专有技术被称为解旋酶依赖性扩增（HDA）。它使用DNA解旋酶在指数扩增期间在65 ℃的恒定温度下分离DNA链。与聚合酶链式反应（PCR）一样，HDA测定使用竞争性内部对照（即，已知浓度的模板DNA掺入到原始样品中），其可以通过与分析物相同的引物扩增，但单独检测;因此允许我们检测直接临床样品中的扩增抑制剂。 阶段I的目标是：1）修改BU微流控装置设计以并入侧向流动条作为使用肉眼检测扩增产物的手段;和2）证明了使用目标1中的器械进行集成测试的可行性，使用在1 mL尿液中以104加标的CT和NG，以建立在改良BU芯片中进行集成测定的概念验证，该芯片包括横向流动带我们在第II阶段的具体目标是开发侧流读数器子系统，将其与BU仪器集成以构建商业前外观/工作原型，简化芯片设计和制造以进行中试规模生产，并在BioHessel/ Quidel实施一次性产品的中试规模生产。在这个阶段，Qiagen是读者最有可能的制造商。在第二阶段结束时，我们应该准备好对新的检测系统进行临床验证。

DESCRIPTION (provided by applicant): This STTR-AT-NIAID seeks to develop an integrated nucleic acid system based on research done by Catherine Klapperich's laboratory at Boston University. The BU lab-on-a-chip includes a micro solid phase extraction (<SPE) column, flap valves and hydrophobic vents to gate fluid movements through micro channels, and multiple reaction chambers for experimental replicates and control reactions. Dr. Klapperich's laboratory has successfully performed nucleic acid amplifications in the chip using BioHelix's isothermal amplification process. Biohelix's proprietary technology is called helicase-dependent amplification (HDA). It uses DNA helicases to separate DNA strands during exponential amplification at a constant temperature of 65:C. Like the polymerase chain reaction (PCR), HDA assays use a competitive internal control (i.e., a template DNA of known concentration spiked into the raw sample) that can be amplified by the same primers as the analyte, but detected separately; hence allowing us to detect amplification inhibitors in direct clinical samples. The objectives of Phase I are to: 1) modify the BU microfluidic device design to incorporate a lateral flow strip as a means of detecting amplification products using the naked eye; and 2) demonstrate the feasibility of performing integrated tests using the device from aim 1 using CT and NG spiked at 104 in 1 mL urine to establish proof of concept for integrated assays performed in a modified BU chip that includes a lateral flow strip. Our specific aims for Phase II will be to develop a lateral flow reader sub-system, integrate it with the BU instrument to build a pre-commercial looks-like / works-like prototypes, simplify chip design and fabrication to allow for pilot scale manufacturing, and implement a pilot scale manufacturing for the disposable at BioHelix / Quidel. At this stage Qiagen is the most likely manufacturer for the reader. At the conclusion of Phase II, we should be ready for clinical validation of the new assay system.