Immunomagnetic Diffractometry for Rapid Biomolecular Detection

用于快速生物分子检测的免疫磁衍射法

• 批准号: 7529962
• 负责人: Cagri Savran
• 金额: $ 18.35万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7529962
• 关键词: American; Antibodies; Area; Behavior; Binding; Biological Assay; Biosensor; Body Fluids; Buffers; Cancer Patient; Cells; Cessation of life; Clinic; Clinical; Complex; Complex Mixtures; Condition; DNA; Decision Making; Detection; Development; Devices; Diagnosis; Dimensions; Disadvantaged; Disease; Doctor of Philosophy; Early Diagnosis; Elements; Event; Figs - dietary; Fluorescence; Folate; Goals; Immunomagnetic Separation; Individual; Label; Laboratory Research; Lasers; Ligands; Lighting; Magnetism; Malignant Neoplasms; Malignant neoplasm of lung; Methods; Microfabrication; Miniaturization; Modeling; Nanotechnology; Nature; Numbers; Operative Surgical Procedures; Optics; Patients; Pattern; Performance; Periodicity; Preparation; Process; Production; Proteins; RNA; Radio; Range; Rate; Reaction; Reaction Time; Refractive Indices; Reporting; Reproducibility; Research Personnel; Sampling; Serum; Signal Transduction; Solid; Solutions; Staging; Students; Surface; System; Techniques; Testing; Theoretical model; Time; Today; Tumor Markers; Universities; Vascular Endothelial Growth Factors; Work; base; biological research; clinically significant; comparative; cost; density; experience; folate-binding protein; high throughput analysis; magnetic beads; mathematical model; model design; nanofabrication; outcome forecast; point of care; portability; prevent; receptor; research study; self assembly; sensor; simulation; size; superparamagnetic beads; tool

DESCRIPTION (provided by applicant): The ability to detect biomolecules in minute quantities is of paramount importance both in biological research and in clinics. Quantitative detection of biomolecules in a specific sample can reveal significant information ranging from cell decision making processes to state of a specific disease. For example, the disease of cancer, that is responsible for half-million deaths in the U.S. every year, can potentially be cured if diagnosed early. One way to perform the diagnosis is to detect tumor markers in body fluids. For a successful early diagnosis, the detection platform must certainly be sensitive and reveal the presence of small amounts of target molecules. The detection platform must also be miniaturizable, quick, relatively simple to operate and inexpensive to enable 1) high- throughput detection and 2) sufficient number of detection platforms in a setting (clinics, research laboratory). Most detection platforms that currently exist require the use of labeling molecules such as fluorescent and radio labels. This increases time, cost and required expertise for operation and renders these label-based platforms not necessarily suitable for miniaturization and high-throughput detection. This proposal covers the development of a biomolecular detection platform that is fluorescence and radio labelfree, simple to fabricate and use, robust and at the same time more sensitive than many existing detection platforms. The proposed system is based on immunomagnetic separation and diffractometry whose combination has not yet been explored. Functionalized magnetic beads are used to capture target molecules from solutions. After this step, the beads (that contain the captured molecules on their surfaces) are simply exposed to a solid surface that is pre-functionalized with secondary receptor molecules. The functionalization is done in an alternating stripe pattern, so that the binding of the beads forms a solid diffraction grating. The illumination of the bound beads forms diffraction modes whose intensities reveal the presence of the beads, and hence the target molecules captured. The detection system does not require any additional amplification steps or fluorescence or radio labels for signaling. The study has 2 PIs and 3 specific aims. Aim 1 is the development of a mathematical model to predict the optical behavior of a bead-based diffraction system. The model will be based on a Monte Carlo simulation and will predict the optimal system parameters (eg. bead size, grating periodicity, surface material, refractive indices) for maximum sensitivity. Aim 2 is the development of the actual setup which will start simultaneously with Aim 1 but later on adapt its results to the experimental setup. Aim 2 will also optimize the system experimentally in terms of receptor and ligand concentrations and background dilutions levels, and explore the limits of detection. Aim 3 is the application of the system to detection of a known tumor marker: vascular endothelial growth factor (VEGF) in complex mixtures such as sera and cell lysate. This aim will explore the "clinical usefulness" of the system with the goal of achieving 500 fM detection sensitivity, a concentration change that distinguishes cancer patients from healthy people.

描述（由申请人提供）：检测微量生物分子的能力在生物学研究和临床中都至关重要。定量检测特定样品中的生物分子可以揭示从细胞决策过程到特定疾病状态的重要信息。例如，每年在美国造成50万人死亡的癌症，如果及早诊断，是有可能治愈的。进行诊断的一种方法是检测体液中的肿瘤标志物。为了成功的早期诊断，检测平台当然必须是敏感的，并揭示少量目标分子的存在。检测平台还必须小型化，快速，相对简单的操作和廉价，以实现1)高通量检测和2)在一个环境中（诊所，研究实验室）足够数量的检测平台。目前存在的大多数检测平台需要使用标记分子，如荧光和无线电标记。这增加了操作的时间、成本和所需的专业知识，并使这些基于标签的平台不一定适合小型化和高通量检测。该提案涵盖了一种生物分子检测平台的开发，该平台无荧光和无线电标记，易于制造和使用，坚固耐用，同时比许多现有的检测平台更敏感。所提出的系统是基于免疫磁分离和衍射，其组合尚未探索。功能化磁珠用于从溶液中捕获目标分子。在这一步之后，这些珠子（表面含有捕获的分子）被简单地暴露在一个固体表面上，这个固体表面被二级受体分子预功能化。功能化是在一个交替的条纹图案中完成的，因此珠子的结合形成一个固体衍射光栅。结合珠子的照明形成衍射模式，其强度显示珠子的存在，从而捕获目标分子。检测系统不需要任何额外的放大步骤或荧光或无线电标记信号。该研究有2个目标和3个具体目标。目标1是建立一个数学模型来预测基于珠的衍射系统的光学行为。该模型将基于蒙特卡罗模拟，并将预测最优系统参数(例如。珠子大小，光栅周期性，表面材料，折射率)的最大灵敏度。目标2是实际装置的发展，它将与目标1同时开始，但稍后将其结果适应实验装置。目的2还将在受体和配体浓度和背景稀释水平方面对系统进行实验优化，并探索检测极限。目标3是应用该系统检测已知的肿瘤标志物：血管内皮生长因子（VEGF）在复杂混合物，如血清和细胞裂解液。这一目标将探索该系统的“临床实用性”，目标是达到500 fM的检测灵敏度，这是区分癌症患者和健康人的浓度变化。