Electromobility Focusing for Separation of Proteins

生物标本（如体液）中蛋白质的分布反映了动物的整体健康状况

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.