Electromobility Focusing for Separation of Proteins

生物标本中的蛋白质分布，如体液，反映了人的整体健康状况 个人。此外，特定标记蛋白的存在或不存在是 表示某些疾病状态，如癌症和心脏病。特异性标志物的测定 复杂生物样品中的蛋白质需要使用极高含量的分析技术 分辨率。蛋白质的大动态范围使典型的分析变得复杂 待分析样品中的浓度，可超过十个数量级。通常，丰富的 蛋白质干扰痕量靶蛋白的检测。这项提议的总体目标是 利用一种称为电迁移率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)，用于分离复杂混合物中的蛋白质。具体的 目的是(1)优化抗蛋白质聚合物整体的孔结构，以最小化 双E成纤维细胞生长因子体系中的流动分散和提供高分辨率的尺寸排除层析 (2)将EFGF和SEC与紫外光检测和质谱学相结合； 以及(3)展示蛋白质分析仪用于目标蛋白质分析的性能。新的 在过去的合同期内发现的聚合物化学物质将被优化并用于 构建用于分离目的蛋白的血管内皮生长因子装置。EFGF的一个主要优势是跟踪 目的蛋白可以被分离和浓缩，同时干扰蛋白可以 被排除在隔离系统之外。新的分析系统将用于检测 前列腺特异性抗原(PSA)是一种游离型和复合型的靶蛋白。

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.