Amplifying Fluorescent Polymer Detection of BW Agents

描述（由申请方提供）：Nomadics将采用扩增荧光聚合物（AFP）技术进行核酸检测。AFP是一类放大由FRET或荧光淬灭事件产生的光学特征的聚合物。将这种固有的信号放大与核酸识别事件所表现出的特异性相结合，将能够实现不依赖于靶扩增或催化的报告物沉积的新一代核酸传感器。这种基于AFP的技术应该对核酸诊断在减小尺寸、重量和功耗方面适应现场操作的需求做出重大贡献。这将使该技术适合用于打击生物恐怖主义。该研究将集中在纳米颗粒AFP检测系统，因为颗粒格式提供了许多优势，在AFP技术的适应PCR分子分析，主要是利用三维信号放大的能力，在固体颗粒，同时保留一个光学透明的流体基质。粒子系统还可以通过单粒子分析来最小化背景并提高灵敏度。此外，AFP组成的变化使我们能够调整珠子的发射颜色;这反过来将使多重分析能够筛选多种试剂。在第一阶段，Nomadics将展示用于核酸检测的APP纳米颗粒技术。这将需要AFP纳米颗粒的制造和表征以及用于附着核酸探针的有效偶联化学的开发。这将使杂交研究能够量化APP纳米颗粒的灵敏度和选择性。在AFP系统的处理和性能的经验将使实验室原型系统的设计和制造。研究将集中在炭疽。针对特定炭疽目标的分子信标的合成和测试将仅在第二阶段进行。第一阶段将包括选择一个适当的炭疽目标的基础上，一个完整的文献检索和设计的分子信标在硅片。为了在第一阶段使用APP纳米颗粒测试这一概念，将使用Nomadics已经设计、合成和测试的另一个目标的探针。第一阶段的目标是证明在低（<10）飞摩尔范围内的高度特异性核酸检测。

DESCRIPTION (provided by applicant): Nomadics will adapt amplifying fluorescent polymer (AFP) technology to nucleic acid detection. AFPs are a class of polymers that amplify optical signatures resulting from FRET or fluorescence quenching events. Combining this inherent signal amplification with the specificity exhibited by nucleic acid recognition events will enable a new generation of nucleic acid sensors that do not rely on target amplification or catalyzed reporter deposition. This AFP-based technology should make a significant contribution to the adaptation of nucleic acid diagnostics towards the demands of field operations in reducing size, weight, and power consumption. This will make the technology suitable for use in countering bioterrorism. The research will focus on nanoparticle AFP detection systems, as the particle format offers a number of advantages in the adaptation of AFP technology to PCR-less molecular analyses-principally, the ability to harness 3-dimensional signal amplification in the solid particle while retaining an optically clear fluid matrix. Particle systems can also minimize background and increase sensitivity through single-particle analyses. Further, variations in the AFP composition allow us to tune the emission color of the bead; this, in turn, will enable multiplex analyses to screen for multiple agents. In Phase I, Nomadics will demonstrate APP nanoparticle technology for the detection of nucleic acids. This will require fabrication and characterization of AFP nanoparticles and the development of efficient coupling chemistries for the attachment of nucleic acid probes. This will enable hybridization studies to quantify the sensitivity and selectivity of APP nanoparticles. Experience in the handling and performance of AFP systems will enable design and fabrication of laboratory prototype systems. Studies will focus on anthrax. Synthesis and testing of a molecular beacon for a specific anthrax targets will only be performed in Phase II. Phase I will include selection of an appropriate anthrax target based on a complete literature search and design of a molecular beacon in silico. In order to test the concept using APP nanoparticles in Phase I, a probe that has already been designed, synthesized, and tested by Nomadics for another target will be used. The goal of Phase I will be to demonstrate highly specific nucleic acid detection in the low (<10) femtomole range.