Isothermal Colorimetric Pathogen DNA Diagnostics

等温比色病原体 DNA 诊断

• 批准号: 7230059
• 负责人: Angelika Niemz
• 金额: $ 21.73万
• 依托单位: KECK GRADUATE INST OF APPLIED LIFE SCIS
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7230059
• 关键词: Benign; Biological Assay; Biological Models; Buffers; Cells; Cerebrospinal Fluid; Class; Classification; Clinical; Condition; Containment; Coupled; Coupling; Cytolysis; DNA; DNA amplification; Detection; Development; Devices; Diagnostic; Drug resistance; Enzymes; Fingerprint; Future; Generations; Genomics; Goals; Gold; Health; Herpesvirus 1; Human; Human Herpesvirus 2; Individual; Infection; Infectious Agent; Lead; Lesion; Life; Methods; Microfluidics; Molecular; Mutation; Nanosphere; Newborn Infant; Numbers; Oligonucleotides; Patients; Preparation; Range; Reaction; Reproducibility; Sampling; Sensitivity and Specificity; Simplexvirus; Site; Spottings; Staging; Swab; System; Validation; Visual; base; clinical application; clinically relevant; improved; internal control; novel; pathogen; point of care; prototype; tool

DESCRIPTION (provided by applicant): The objective of this proposal is to develop a rapid, sensitive DMA diagnostic assay, which combines a novel isothermal DNA amplification reaction (EXPAR) with colorimetric detection through DNA-functionalized gold nanospheres in a format applicable to point-of-care settings. The EXPAR reaction amplifies short trigger oligonucleotides >10A6 fold at 55¿C in a matter of minutes. Detection through the aggregation of DNA- functionalized gold nanospheres provides a visual readout in the form of a red or blue spot. We will develop the assay to detect Herpes Simplex Virus type 1 and 2 (HSV1 and HSV2), which, although generally benign, can lead to life-threatening infections in newborns and immuno-compromised individuals. The widespread occurrence of HSV and the emergence of drug-resistant strains necessitate the development of diagnostic tools to facilitate effective treatment of patients and to contain further spread. The long-term goal will be to expand this assay format to other classes of pathogens. The specific aims of the proposed project are to: (1) Develop methods for generating trigger oligonucleotides from HSV genomic target DNA, which will enable specific identification of HSV1 and/or HSV2, as well as identification of drug-resistance mutations. (2) Optimize the amplification of trigger oligonucleotides coupled to detection via DNA nanosphere aggregation, in single or multiplexed format, with the appropriate sensitivity, selectivity and robustness required for clinical applications. (3) Optimize and validate complete assays that combine trigger generation with trigger amplification and detection to enable rapid, sensitive, specific, robust, and reproducible detection of HSV1, HSV2, and drug resistance mutations thereof. This proposal focuses on fully developing and optimizing this novel assay, which we plan to implement in a micro fluidic device in a subsequent effort. The proposed project will enable the development of new systems for DNA-based molecular diagnostics in point of care settings, which is expected to facilitate rapid identification and containment of infectious agents, and thus have a significant impact on human health.

描述（由申请人提供）：这是通过以DNA官能化的金纳米的比色检测来开发快速，敏感的DMA诊断测定的热DNA扩增反应（Expar）反应放大短触发寡核苷酸> 10a6折叠55¿ c在几分钟内。良性可以导致新生儿促进性的人的生命线感染，而耐药菌株的出现是诊断工具的开发，以促进治疗方法，并通过扩展到其他扩展。病原体的类别。临床应用的等式。在随后的努力中，完全开发和优化这是一个次云母设备中的微流体装置。因此，对人类健康有重大影响。