| Summary: | <p>Abstract</p> <p>Background</p> <p>Low-abundance proteins are difficultly observed on the two-dimensional gel electrophoresis (2-DE) maps of urine proteome, because they are usually obscured by high-abundance proteins such as albumin and immunoglobulin. In this study, a novel fractionation method was developed for enriching low-abundance proteins by removing high-abundance proteins and progressive elution with salts of various concentrations.</p> <p>Results</p> <p>Stepwise weak anion exchange (WAX) chromatography, which applied DEAE-Sephacel resin with non-fixed volume elution, was used to fractionate urine proteome prior to performing 2-DE. Urine proteome was separated into four fractions by progressively eluting the column with 0 M, 50 mM, 100 mM, and 1 M NaCl solutions. Most of the heavy and light immunoglobulin chains appeared in the eluent. After the high-abundance proteins were removed, various low-abundance proteins were enriched and could be easily identified. The potential of this method for obtaining diversified fractionations was demonstrated by eluting the column separately with Na<sub>2</sub>SO<sub>4 </sub>and MgCl<sub>2 </sub>solutions. The 2-DE maps of the fractions eluted with these different salt solutions of identical ionic strength revealed markedly different stain patterns.</p> <p>Conclusion</p> <p>The present study demonstrated that this fractionation method could be applied for purposes of enriching low-abundance proteins and obtaining diversified fractionations of urine, and potentially other proteomes.</p>
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