Development of DNA Logic Operations for Viral Diagnostics

描述（由申请人提供）：抗生素的过度使用导致美国抗生素耐药率飙升，更多的抗生素处方用于上呼吸道感染，如呼吸道合胞病毒（RSV），而不是其他任何适应症。医生自述这些处方不适用于病毒诊断。快速诊断测试将为医生提供避免不当使用抗生素所需的信息。由于其独特的特异性和可塑性，基于抗体的策略仍然是许多病毒检测平台的关键组成部分。然而，基于高灵敏度抗体的检测技术的一个主要障碍是抗体和非靶标之间固有的非特异性相互作用。本应用程序描述了一种结合DNA报告标记和DNA-抗体标记之间的逻辑操作的方法，以开发基于DNA标记的病毒检测的新范式。在提出的方法中，不是指定唯一的，但任意选择的DNA序列作为DNA-抗体标签，而是故意设计DNA序列以方便标签之间的逻辑操作。在目标1中，这些“DNA逻辑标签”将被设计、测试，并与金纳米颗粒表面的抗体共偶联。据推测，这种逻辑标签方法将允许使用逻辑非操作来控制非特异性抗体相互作用（目标2），并允许使用逻辑或操作来独立微调检测灵敏度和特异性（目标3）。这些假设将通过测量逻辑操作对RSV检测分析在日益复杂的生物环境中进行的敏感性和特异性的影响来进行评估。高灵敏度和特异性的诊断工具，如本文提出的，将在即时诊断中变得越来越重要，以快速识别病原体并帮助指导选择适当的治疗方法。

DESCRIPTION (provided by applicant): The overuse of antibiotics has led to soaring rates of antibiotic resistance in the U.S. More antibiotic prescriptions are written for upper respiratory infections, such as respiratory syncytial virus (RSV), than for any other indication. Physicians self-report that these prescriptions would be inappropriate for a viral diagnosis. Rapid diagnostic tests would give physicians the information needed to avoid inappropriate antibiotic use. Because of their unique specificity and plasticity, antibody-based strategies continue to be a key component of many virus detection platforms. However, a major stumbling block of high-sensitivity antibody-based detection technologies is the innate presence of non-specific interactions between antibodies and non-targets. This application describes a method to combine DNA reporter tags and logical operations among DNA-antibody tags to develop a new paradigm in DNA tag-based virus detection. In the proposed approach, instead of specifying unique, but arbitrarily chosen DNA sequences as DNA-antibody tags, the DNA sequences are purposely designed to facilitate logical operations among tags. In Aim 1 these "DNA logic tags" will be designed, tested, and co-coupled with antibodies to gold nanoparticle surfaces. It is hypothesized that this logic tag approach will permit control of non-specific antibody interactions using a logical NOT operation (Aim 2), and a means to independently fine-tune detection sensitivity and specificity using logical OR and operations among tags (Aim 3). These hypotheses will be evaluated by measurements of the effect of the logical operations on the sensitivity and specificity of an RSV detection assay performed in an increasingly complex biological environment. Highly sensitive and specific diagnostic tools, such as proposed here, will become increasingly important in point-of-care diagnostics to rapidly identify pathogens and help guide the choice of appropriate therapeutic treatment. PUBLIC HEALTH RELEVANCE: More antibiotic prescriptions are written for upper respiratory infections (an estimated 100 million prescriptions a year) than for any other indication, and doctors self-report that these prescriptions would be inappropriate if a viral diagnosis was available. Rapid, simple and accurate diagnostic tests for early detection of respiratory virus infections are sorely needed. The goal of this project is to evaluate a new viral detection technology for the medically important exemplar, respiratory syncytial virus.