Optical Encoding Technology for Viral Screening Panels

用于病毒筛查面板的光学编码技术

• 批准号: 7747747
• 负责人: ROBERT C HAUSHALTER
• 金额: $ 7.09万
• 依托单位: PARALLEL SYNTHESIS TECHNOLOGIES, INC.
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-15 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7747747
• 关键词: Address; Animals; Arts; Bacteria; Bioinformatics; Biological; Biological Assay; Biotin; Code; Collaborations; Complementary DNA; Computer software; DNA; Data; Detection; Development; Devices; Environment; Equipment; Family; Figs - dietary; Generations; Goals; Human; Information Systems; Ions; Label; Laboratories; Laboratory Scientists; Lasers; Location; Measurement; Measures; Methods; Methyl Green; Nucleic Acids; One-Step dentin bonding system; Optics; Pathogen detection; Performance; Phase; Phycoerythrin; Plants; Process; Protocols documentation; RNA; Reaction; Reader; Reading; Relative (related person); Reproducibility; Reverse Transcription; Running; Sampling; Scanning; Scientist; Screening procedure; Series; Silicon; Slide; Source; Speed; Streptavidin; Suspension substance; Suspensions; System; Technology; Testing; Text; Vial device; Viral; Viral Genome; Virus; Writing; base; biothreat; cost; design; detector; influenzavirus; innovation; laboratory facility; novel; pathogen; portability; prevent; public health relevance; research study; standard measure; viral RNA; viral detection

DESCRIPTION (provided by applicant): The last decade has witnessed substantial improvement in the development of parallel screening technologies designed to detect many types of biological threats. One type of screening technology has concentrated on mitigating threats caused by viral Biothreat agents by using multiplex PCR followed by hybridization onto optically encoded beads from Luminex. However, the rather small number of usable bead optical codes (~30-40), and the delicate flow cytometer based detector, prevent the detection of all viral threats in one panel or use of the screen outside of a controlled laboratory environment. To address these issues, a system based on Parallume (www.parallume.com) optical encoding, which can support thousands of optical codes, will be used to encode beads that will be read by a completely portable, battery-powered reader. The final goal will be to develop a screening panel capable of detecting, differentiating and identifying all known viral families in any location by using a portable detection system. In collaboration with scientists at Lawrence Livermore National Laboratory (LLNL), Parallel Synthesis Technologies, Inc. (PSTI) will formulate a Parallume-encoded bead set containing 15 optical codes which will be used in a direct comparison against a 15-bead Luminex-based Viral Screening Panel (VSP) already in use at LLNL to detect human influenza viruses. Using biotinylated primers designed by the LLNL Bioinformatics Group, PCR for 15 regions of viral genomes are performed in a single reaction. A target region of each PCR product contains a sequence which is hybridized to a capture probe on the optically encoded bead. By treating the PCR product after hybridization with a streptavidin-phycoerythrin (SAPE) conjugate to label the biotinylated primers, the presence of the viral genome is detected by a serially diluted, threshold SAPE level on an encoded bead as compared to the controls. In the Phase I effort, Parallel and LLNL will compare the state-of-the-art Luminex system with the Parallume encoding technology by characterizing the same reactions on a divided PCR product mixture. The Parallel samples will be characterized using Parallume beads and Parallel's robust Multiplex Assay Reader System (MARS), which has replaced the fragile flow cytometer and lasers with micromachined silicon bead localization slides and super-bright signage LEDs, and the measurement rate, accuracy, precision and cost of the two systems directly compared. The combination of the Parallel and LLNL technologies could allow all known viruses to be screened anywhere in one PCR reaction. PUBLIC HEALTH RELEVANCE This innovation seeks to create a viral screening panel (VSP) whereby a human, animal, or plant sample containing DNA is subjected to a simultaneous screen for multiple pathogens (viruses in the current application) within a single vial. Limited virus detection technologies exist, but are hampered by technical limitations not possessed by the proposed VSP. This novel VSP proposes a method for expanding the number of viruses currently detectable in a single screen and adds to the potential functionality of the panel by being able to incorporate other screens (e.g., bacterial and other non-viral pathogens) into this single, multiplexed panel. This technology also offers a dramatically less-expensive reader (VSP analysis equipment) to accompany this new product.

描述（由申请人提供）：最近十年见证了旨在检测​​许多类型的生物威胁的平行筛查技术的发展。一种类型的筛查技术集中在通过使用多重PCR，然后将杂交在Luminex的光学编码珠上进行杂交引起的缓解威胁。但是，相当少数可用的珠光学代码（〜30-40）以及基于流式细胞仪的精致检测器，可以防止在一个面板中检测所有病毒威胁或在受控实验室环境外的屏幕上使用屏幕。为了解决这些问题，将使用基于差异（www.parallume.com）光学编码的系统来支持数千个光学代码，该系统将用于编码由完全便携式，电池供电的读取器读取的珠子。最终目标是开发一个能够使用便携式检测系统在任何位置检测，区分和识别所有已知病毒家族的筛选面板。 与Lawrence Livermore国家实验室（LLNL）的科学家合作，并行合成技术，Inc。（PSTI）将制定一个由偏见编码的珠子组，其中包含15种光学代码，这些珠子将用于与15-Bead Luminex的直接比较，该光学代码与15-Bead Luminex的病毒筛选面板（VSP）（在LLNL上都具有LALNL的使用）来检测。使用LLNL生物信息学组设计的生物素化引物，在单个反应中进行了15个病毒基因组区域的PCR。每个PCR产物的目标区域包含一个序列，该序列与光学编码珠上的捕获探针杂交。通过与链霉亲和素 - 磷酸（SAPE）偶联物杂交后处理PCR产物，以标记生物素化引物，与对照组相比，通过在编码的珠子上的串行稀释的阈值SAPE水平检测到病毒基因组的存在。在第一阶段的工作中，平行和LLNL将通过表征分开的PCR产品混合物上相同的反应来比较最先进的Luminex系统与编码技术的编码技术。平行样品将使用拼奔珠和平行的鲁棒多路复用测定读取器系统（MARS）来表征，该系统已用微机械硅珠定位幻灯片和超级黑色标志LED和测量率，准确性，准确性，精确性和成本直接比较，取代了脆弱的流式细胞仪和激光器。平行技术和LLNL技术的组合可以使所有已知病毒在一个PCR反应中的任何位置进行筛查。公共卫生相关性这项创新旨在创建病毒筛查面板（VSP），从而使包含DNA的人类，动物或植物样品同时进行单个小瓶中多种病原体（当前应用中的病毒）的同时筛查。存在有限的病毒检测技术，但由于所提出的VSP所没有的技术局限性受到阻碍。这部新颖的VSP提出了一种方法，可以通过将其他筛选（例如细菌和其他非病毒病原体）纳入该单个单个多路面板，将当前可检测到的病毒数量扩展到面板的潜在功能。这项技术还提供了较低的读者（VSP分析设备），以伴随着这一新产品。