Effect of immunoaffinity depletion of human serum during proteomic investigations

Anastasia K. Yocum, Kenneth Yu, Tomoyuki Oe, Ian A. Blair

Research output: Contribution to journalArticlepeer-review

101 Citations (Scopus)

Abstract

Controversy exists regarding the proper mining of the human serum proteome. Because of the analytical challenges of accurately measuring samples containing a very large dynamic range of protein concentrations, current practices have employed depletion of the highly abundant housekeeping serum proteins, such as albumin and immunoglobins. There is question as to the selectivity of depletion, namely, is there loss of other non abundant serum proteins along with albumin, haptoglobin and other commonly depleted proteins. In this study, human serum was analyzed with and without immunoaffinity depletion of the six most abundant proteins by multidimensional liquid chromatography tandem mass spectrometry. Two replicates of each experiment were conducted and compared against one another. In both depleted and nondepleted replicates there was a 73% and 72% overlap of identified peptides and a 64% and 78% overlap of identified proteins, respectively. Of 262 unique proteins identified in the four experiments, 82 were found in common to all four experiments, 142 unique to the depleted serum, and 38 unique to the nondepleted serum. Although serum depletion of highly abundant proteins significantly increased the number of proteins identified, both the degree of sample complexity and this depletion method resulted in a nonselective loss of other proteins.

Original languageEnglish
Pages (from-to)1722-1731
Number of pages10
JournalJournal of Proteome Research
Volume4
Issue number5
DOIs
Publication statusPublished - 2005 Sep
Externally publishedYes

Keywords

  • 2D-LC-MS/MS
  • Albumin-depletion
  • Biomarker
  • INTERACT
  • Mass spectrometry
  • ProteinProphet/Peptide Prophet
  • Proteomics
  • Serum

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

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