Electromobility Focusing for Separation of Proteins

用于蛋白质分离的电动聚焦

• 批准号: 7013144
• 负责人: MILTON L. LEE
• 金额: $ 35万
• 依托单位: BRIGHAM YOUNG UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-02-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7013144
• 关键词: alpha fetoprotein; biomarker; blood chemistry; blood proteins; carcinoembryonal antigen; chorionic gonadotropin; clinical research; electrofocusing; human tissue; prostate specific antigen; protein purification; serology /serodiagnosis; technology /technique development; thymidine kinase

Protein distributions in biological specimens, such as body fluids, are reflective of the overall health of individuals. In addition, the presence or absence of specific marker proteins is indicative of certain disease states, such as cancer and heart disease. Determination of specific marker proteins in complex biological samples requires the use of analytical techniques with extremely high resolving power. Typical analyses are complicated by the wide dynamic range of protein concentrations in samples to be analyzed, which can exceed ten orders of magnitude. Often, abundant proteins interfere with the detection of trace target proteins. The overall objective of this proposal is to exploit the unique properties of a relatively new analytical technique called electromobility focusing or electric field gradient focusing (EFGF) for separating proteins in complex mixtures. The specific aims are (1) to optimize the pore structure of a protein-resistant polymeric monolith for minimizing flow dispersion in a dual-EFGF system and providing high resolution size exclusion chromatography (SEC) of proteins; (2) to serially integrate EFGF and SEC with UV detection and mass spectrometry; and (3) to demonstrate the performance of the protein analyzer for target protein analysis. New polymer chemistries discovered during the past contract period will be optimized and utilized for constructing EFGF devices for isolation of target proteins. A major advantage of EFGF is that trace target proteins can be isolated and concentrated while, at the same time, interfering proteins can be excluded from the separation system. The new analytical system will be demonstrated for detection of prostate specific antigen (PSA) as a target protein in its free and complexed forms.

生物标本中的蛋白质分布，例如体液，反映了 个人。另外，存在或不存在特定标记蛋白 指示某些疾病状态，例如癌症和心脏病。确定特定标记 复杂生物样品中的蛋白质需要使用极高的分析技术 解决力量。典型的分析因蛋白质的广泛动态范围而变得复杂 要分析的样品中的浓度，可能超过十个数量级。通常，丰富 蛋白质干扰痕量靶蛋白的检测。该提议的总体目的是 利用一种称为电动性聚焦的相对较新的分析技术的独特特性 或电场梯度聚焦（EFGF），用于在复杂混合物中分离蛋白质。具体 目的是（1）优化抗蛋白质的聚合物整体的孔结构，以最大程度地减少 双EFGF系统中的流量分散，并提供高分辨率尺寸排除色谱法 （秒）蛋白质； （2）用UV检测和质谱法串行整合EFGF和SEC； （3）证明蛋白质分析仪的性能进行靶蛋白分析。新的 在过去的合同期间发现的聚合物化学成分将被优化和利用 构建用于隔离靶蛋白的EFGF设备。 EFGF的主要优势是该痕迹 靶蛋白可以分离和浓缩，同时可以干扰蛋白 排除在分离系统之外。将展示新的分析系统以检测 前列腺特异性抗原（PSA）作为靶蛋白的游离形式。