Novel, single-molecule aptamer selection method

新颖的单分子适体选择方法

• 批准号: 6687228
• 负责人: MARTIN GUTHOLD
• 金额: $ 11.37万
• 依托单位: NANOMEDICA, INC.
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-06 至 2005-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6687228
• 关键词: analytical method; atomic force microscopy; fluorescence microscopy; image enhancement; light microscopy; molecular shape; nanotechnology; nucleic acid quantitation /detection; oligonucleotides; technology /technique development

DESCRIPTION (provided by applicant): Molecular recognition is the underlying principle in many biomedical applications, such as labeling and imaging specific proteins, organelles, cells, tissues and organs, detecting and quantifying clinical analytes, and discovering target-specific drug candidates. Currently, the primary technology for identifying research and diagnostic ligands is monoclonal antibody selection by hybridoma screening. Aptamers - an emerging class of antibody mimetics - have been used to a much lesser extent but are gaining attention in academic circles. Aptamers are short oligonucleotides that fold into target-specific 3D structures. Although aptamers have many potential advantages over antibodies, their adoption has been slow, because 1) many researchers presume that SELEX is the only way to select aptamers, 2) SELEX is regarded by many as a cumbersome, labor-intensive and time-consuming method that requires specialized expertise, and 3) SELEX is heavily protected by patents that are aggressively enforced. The long-term goal of this proposal is to develop a convenient and fully automated instrument-based system for rapid selection of aptamers useful in the molecular analysis of cancer. NanoMedica plans to commercialize an affordable, modular system that will empower bench scientists to select and characterize high-affinity aptamers for studying cancer, signal transduction, and protein-protein interactions. If successful, this system will be fast, efficient and much more amenable to broad dissemination than SELEX. At the core of this technique is unique instrumentation that combines a nanoManipulator-Atomic Force Microscope (nM-AFM) with an inverted optical microscope to achieve fast imaging, high resolution and single-molecule detection and manipulation capabilities. The instrument will be able to detect single aptamers specifically bound to immobilized target molecules by fluorescence microscopy. The fluorescence signal will be used to guide the AFM tip towards the bound aptamer to enable single-molecule detection and aptamer extraction. Before extraction, the aptamer's affinity for its target will be measured in situ by dynamic force spectroscopy. After extraction, the aptamer molecule will be amplified by PCR and further characterized by traditional biochemical methods.

描述（由申请人提供）：