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Microfluidic Selection of Aptamers for Biological Purification Applications

用于生物纯化应用的适体的微流体选择

基本信息

批准号：
8011309
负责人：
Qiao Lin
金额：
$ 19.56万
依托单位：
COLUMBIA UNIV NEW YORK MORNINGSIDE
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-01-01 至 2012-12-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Microfluidic Selection of Aptamers for Biological Purification Applications Principal Investigator: Qiao Lin, Columbia University Aptamers are oligonucleotides that bind to small molecules or proteins. Aptamers are isolated through an in vitro selection and amplification procedure called systematic evolution of ligands by exponential enrichment (SELEX). The procedure is based on an affinity selection, followed by amplification of nucleic acids that are "binders" from large random libraries. Aptamers can be developed for an extremely broad spectrum of analytes with high affinity, can possess well controlled target selectivity, and can be synthesized to bind targets with predefined characteristics. In particular, aptamer binding in general exhibits strong temperature dependence; thus, aptamers may specifically bind target analytes at a predefined temperature and reversibly decouple from the targets at a modestly different, yet also predefined, temperature. This property is attractive to biomedical applications such as affinity purification, as it can enable specific purification with thermally activated release and isocratic elution of analytes. Conventional SELEX instruments have enabled great progress in aptamers, but their use is generally labor-intensive and time-consuming. These limitations can be addressed by leveraging microfluidic technology. We propose to pursue proof-of-concept demonstration of a microfluidic SELEX system that integrates all steps of the SELEX method to allow automated development of aptamers with predefined temperature-dependent binding characteristics for applications to affinity purification of analytes. Our specific aims include: (1) developing a bead-based polymerase chain reaction (PCR) technique in a microchannel to establish its applicability to microfluidic SELEX; (2) integrating microchip DNA selection and amplification to create a prototype microfluidic SELEX system; and (3) characterize and validate the prototype system by using it to select aptamers against well established proteins. The proposed research is relevant to public health because it will provide a platform for selecting aptamers for affinity purification of analytes such as proteins, small molecules and cells involved in biomedical research on diseases. In addition, this system could also be used to develop aptamers for applications to target validation, drug discovery, diagnostics, and therapy. PUBLIC HEALTH RELEVANCE (provided by applicant): Microfluidic Selection of Aptamers for Biological Purification Applications Principal Investigator: Qiao Lin, Columbia University This research will pursue proof-of-concept demonstration of a microfluidic system that integrates all steps of the method of systematic evolution of ligands by exponential enrichment (SELEX). The system will allow automated development of aptamers with predefined temperature-dependent binding characteristics for analyte purification applications. It is relevant to public health because it will provide a platform for selecting aptamers for affinity purification of analytes such as proteins, small molecules and cells involved in biomedical research on diseases. In addition, this system could also be used to develop aptamers for applications to target validation, drug discovery, diagnostics, and therapy.

描述（由申请人提供）：用于生物纯化应用的适体的微流体选择主要研究者：乔琳，哥伦比亚大学适体是与小分子或蛋白质结合的寡核苷酸。通过体外选择和扩增程序分离适体，该程序称为指数富集配体系统进化（SELEX）。该程序是基于亲和选择，然后从大的随机文库扩增作为“结合剂”的核酸。适体可以被开发用于具有高亲和力的极其广谱的分析物，可以具有良好控制的靶标选择性，并且可以被合成以结合具有预定义特征的靶标。特别地，适体结合通常表现出强烈的温度依赖性;因此，适体可以在预定义的温度下特异性地结合靶分析物，并且在适度不同但也是预定义的温度下可逆地与靶分离。这种性质对于生物医学应用如亲和纯化是有吸引力的，因为它可以通过分析物的热活化释放和等度洗脱实现特异性纯化。传统的SELEX仪器已经在适体方面取得了很大的进展，但它们的使用通常是劳动密集型和耗时的。这些限制可以通过利用微流体技术来解决。我们建议追求的概念验证演示的微流控SELEX系统，集成的SELEX方法的所有步骤，以允许自动化开发的适体与预定义的温度依赖性的结合特性的应用程序的亲和纯化的分析物。我们的具体目标包括：（1）在微通道中开发基于珠粒的聚合酶链式反应（PCR）技术以建立其对微流体SELEX的适用性;（2）整合微芯片DNA选择和扩增以创建原型微流体SELEX系统;以及（3）通过使用原型系统来选择针对良好建立的蛋白质的适体来表征和验证原型系统。拟议的研究与公共卫生有关，因为它将提供一个平台，用于选择适体，用于亲和纯化疾病生物医学研究中涉及的蛋白质，小分子和细胞等分析物。此外，该系统还可用于开发应用于靶向验证、药物发现、诊断和治疗的适体。公共卫生相关性（由申请人提供）：用于生物纯化应用的适体的微流体选择主要研究者：乔琳，哥伦比亚大学本研究将对集成了指数富集配体系统进化方法（SELEX）的所有步骤的微流体系统进行概念验证。该系统将允许自动开发具有预定义温度依赖性结合特征的适体，用于分析物纯化应用。它与公共卫生有关，因为它将提供一个平台，用于选择适体，用于亲和纯化疾病生物医学研究中涉及的蛋白质，小分子和细胞等分析物。此外，该系统还可用于开发应用于靶向验证、药物发现、诊断和治疗的适体。

项目成果

期刊论文数量（15）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Microcantilever-Based Label-Free Characterization of Temperature-Dependent Biomolecular Affinity Binding.

DOI：
10.1016/j.snb.2012.02.045
发表时间：
2013-01-01
期刊：
SENSORS AND ACTUATORS B-CHEMICAL
影响因子：
8.4
作者：
Wang, Bin;Huang, Fengliang;ThaiHuu Nguyen;Xu, Yong;Lin, Qiao
通讯作者：
Lin, Qiao

Surface-enhanced Raman spectroscopy based quantitative bioassay on aptamer-functionalized nanopillars using large-area Raman mapping.

DOI：
10.1021/nn401199k
发表时间：
2013-06-25
期刊：
ACS NANO
影响因子：
17.1
作者：
Yang, Jaeyoung;Palla, Mirko;Bosco, Filippo Giacomo;Rindzevicius, Tomas;Alstrom, Tommy Sonne;Schmidt, Michael Stenbaek;Boisen, Anja;Ju, Jingyue;Lin, Qiao
通讯作者：
Lin, Qiao

A MEMS-Based Approach to Single Nucleotide Polymorphism Genotyping.