RAPID: Development of a handheld, rapid molecular diagnostic tool for Ebola

RAPID：开发埃博拉手持式快速分子诊断工具

• 批准号: 1511093
• 负责人: David Galbraith
• 金额: $ 20万
• 依托单位: University of Arizona
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-01-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1511093&HistoricalAwards=false
• 关键词: RAPID; Development; handheld; rapid; molecular

1511093 Galbraith, David W.University of Arizona Ebola virus infection provides considerable current cause for concern due to a lack of effective interventions, its high lethality, and a lack of rapid and sensitive means to detect viral presence. Molecular diagnostics, based on detection of the Ebola virus genome using the Polymerase Chain Reaction (PCR) technique developed in the US by Nobel Laureate Cary Mullis, represents a sensitive approach to screen for infection. However, current PCR instruments are large, expensive and complicated, and do not operate at the speeds required to make clinical decisions. The investigators have developed a novel PCR device, termed Droplet-On-Thermocouple Silhouette quantitative PCR (DOTS qPCR), which operates on the basis of sensing interfacial effects instead of the established fluorescence detection of the PCR reaction. This innovative detection method enables much faster analysis times with sample-to-answer times within 5 minutes. This RAPID project aims to further develop the device to demonstrate its applicability for detection of Ebola virus from human samples, to interface the device with standard smartphones for data collection, processing and transmission, and to show how it can be used as a handheld device. Beyond the rapid detection of Ebola virus, the DOTS-PCR device will be universally applicable for detection of any biological disease organism of concern to the US National health, agriculture, and security. The novel PCR methodology (DOTS qPCR) utilizes for the first time innovative engineering principles on interfaces to achieve droplet actuation, inhibition relief, and sensing of the PCR reaction, resulting in sample-to-answer times as short as 5 minutes. Towards diagnosis of the presence of Ebola virus, the investigators propose to demonstrate reproducibility, differentiation of virus species, sub-picogram limit of detection, and thermocycling speeds of 28 s/cycle in the presence of blood/tissue contaminants. The specific aims of this project are (i) development of a second-generation handheld DOTS qPCR appropriate for diagnostic use, (ii) demonstration of the ability to identify target in the presence of typical blood/tissue contaminants, (iii) integration of smartphone-based identification of amplicons using the interfacial effect instead of traditional fluorescence sensing, with development of appropriate software for data processing and result communication. Although aimed at detection of Ebola, this technology clearly has general applicability in the high-throughput, low-cost detection of other specific biological organisms and agents. The intellectual merit underpinning these aims rests in the innovative exploitation of interfacial effects on the surface of aqueous reaction droplets and the surrounding oil phase associated with the partitioning of contaminating proteins and the formation of amplification products resulting in detection speeds far superior to traditional, fluorescence based instruments.

1511093 Arizona Ebola病毒感染的Galbraith，David W. University，由于缺乏有效的干预措施，高致死性以及缺乏快速而敏感的手段来检测病毒存在，这为当前的原因提供了相当大的当前原因。 基于诺贝尔奖获得者Cary Mullis在美国开发的聚合酶链反应（PCR）技术对埃博拉病毒基因组检测的分子诊断，代表了一种敏感的感染方法。但是，当前的PCR仪器很大，昂贵且复杂，并且不以做出临床决策所需的速度运行。 研究人员开发了一种新型的PCR设备，称为液滴式轮廓剪影定量PCR（DOTS QPCR），该设备是基于感应界面效应而不是确定的PCR反应荧光检测的。这种创新的检测方法可以在5分钟内使用样本到撤离时间更快地分析时间。这个快速的项目旨在进一步开发该设备，以证明其可用于检测埃博拉病毒从人类样品中的适用性，以将设备与标准智能手机连接到数据收集，处理和传输，并展示如何将其用作手持设备。除了快速检测到埃博拉病毒外，DOTS-PCR设备还普遍适用于对美国国家健康，农业和安全的任何生物疾病生物的发现。 新型的PCR方法（DOTS QPCR）首次在界面上使用创新的工程原理，以实现液滴致动，抑制作用和对PCR反应的传感，从而导致样本之间短达5分钟。为了诊断埃博拉病毒的存在，研究人员建议在存在血液/组织污染物的情况下证明可重复性，病毒物种的分化，检测的亚微图限和28 s/cycle的热循环速度。该项目的具体目的是（i）开发适合诊断使用的第二代手持点QPCR，（ii）证明在存在典型的血/组织污染物的情况下识别目标的能力，（iii）基于智能手机的识别基于智能手机的鉴定，使用插入式效果而不是传统荧光效应而不是传统的荧光效应，而不是传统的荧光效应，而不是使用传统的荧光效应来处理适用于数据处理和数据处理。尽管目的是检测埃博拉病毒，但该技术显然在对其他特定生物生物体和剂的高术，低成本检测中具有一般适用性。这些目标的基础的智力优点取决于对界面对水反应液滴表面的界面作用的创新剥削，以及与污染蛋白的分配相关的周围油相，并形成扩增产物的形成，导致检测速度与传统的，荧光基于荧光的仪器相比较高。