Depletion of the high-abundance plasma proteins

Abstract

Body fluids, like plasma and urine, are comparatively easy to obtain and are useful for the detection of novel diagnostic markers by applying new technologies, like proteomics. However, in plasma, several high-abundance proteins are dominant and repress the signals of the lower-abundance proteins, which then become undetectable either by two-dimensional gels or chromatography. Therefore, depletion of the abundant proteins is a prerequisite for the detection of the low-abundance components. We applied affinity chromatography on blue matrix and Protein G and removed the most abundant human plasma proteins, albumin and the immunoglobulin chains. The plasma proteins, prior to albumin and immunoglobulin depletion, as well the eluates from the two chromatography steps were analyzed by two-dimensional electrophoresis and the proteins were identified by MALDI-TOF-MS. The analysis resulted in the identification of 83 different gene products in the untreated plasma. Removal of the high-abundance proteins resulted in the visualization of new protein signals. In the eluate of the two affinity steps, mostly albumin and immunoglobulin spots were detected but also spots representing several other abundant plasma proteins. The methodology is easy to perform and is useful as a first step in the detection of diagnostic markers in body fluids by applying proteomics technologies.

Human body fluid proteome analysis

“…However, the binding of albumin to Cibacron blue dyes is nonspecific, and the sensitivity and specificity are not as effective as mAb-based immunoaffinity resin or columns [15][16][17][18]. Removal of IgG can be realized with Protein G resins or columns [15][16][17][18][19][20]. Comparing to Cibacron blue dye and Protein G methods, immunoaffinity depletion using multiple affinity removal columns (MARC) is more effective because it can simultaneously remove multiple abundant proteins, with minimal carryover, high longevity, and minimal nonspecific binding [16,17,21,22].…”

Recent advances in blood‐related proteomics

“…Intact as well as partially degraded proteins or protein fragments circulate in the blood. Furthermore, genetic polymorphisms as well as numerous post-translationally modified forms of proteins are present, [17][18][19][20]. Removal of fibrinogen (Factor I) is easily obtained by clotting.…”

Interaction of serum albumin with vinyl sulfonate azo dye

“…Effective removal of high-abundance plasma proteins is of high importance in proteomics studies, which are aimed to identify biomarkers of diseases [1]. To separate the albumin, dye-affinity chromatography is commonly used, and Cibacron blue F3G-A (CB) is a ligand of high popularity [2].…”