Effect of Plasma Protein Depletion on BNP-32 Recovery

Abstract

Depletion of abundant proteins from plasma and serum is an important initial step in many biomarker discovery platforms(1). Decreasing the concentrations of highly abundant proteins (e.g., albumin, IgG, and antitrypsin) facilitates the use of contemporary proteomics technologies, such as gel electrophoresis and mass spectrometry, for detection and identification of low-abundant proteins. Furthermore, decreasing abundant protein concentrations may also improve immunoprecipitation recovery efficiencies for targeted low-abundant species by decreasing nonspecific binding (i.e., shielding the antigen-binding domain) to the antibody and/or solid supports. A notable pitfall to depletion strategies is the potential for unintentionally removing low-abundant plasma or serum proteins. These low-abundant species may be bound specifically or nonspecifically to the depletion ligand, depletion target protein (e.g., carrier proteins), or the solid support(s). Thus, it is important to critically evaluate the effectiveness of abundant plasma protein depletion for enhancing the study of low-abundant protein biomarker(s). We have been actively developing a targeted biomarker discovery platform for characterizing the circulating forms of B-type natriuretic peptide (BNP) that includes protein depletion strategies, immunoprecipitation, gel electrophoresis, isotope dilution (absolute quantification), and nanoflow liquid chromatography coupled to high-performance hybrid Fourier transform mass …