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DIGE Analysis of Immunodepleted Plasma.

Paul Dowling1, Kay Ohlendieck2

  • 1Department of Biology, Maynooth University, National University of Ireland, Maynooth, Co. Kildare, Ireland. paul.dowling@mu.ie.

Methods in Molecular Biology (Clifton, N.J.)
|November 15, 2022
PubMed
Summary

High-abundance protein depletion from plasma using immunodepletion enhances the detection of low-abundance proteins. This method, combined with 2D-DIGE, aids in discovering disease biomarkers.

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2D-DIGE proteomics.

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Area of Science:

  • Proteomics
  • Biomarker Discovery
  • Analytical Chemistry

Background:

  • Biofluid proteomes exhibit vast dynamic ranges, complicating the analysis of low-abundance proteins.
  • High-abundance proteins can mask the detection of medium and low-abundance proteins in techniques like 2D-DIGE.
  • Identifying disease-specific biomarkers requires sensitive detection of low-abundance proteins.

Purpose of the Study:

  • To describe the upstream immunodepletion of high-abundance proteins from plasma samples.
  • To detail the subsequent analysis using fluorescence two-dimensional difference gel electrophoresis (2D-DIGE).
  • To highlight the combined utility of immunodepletion and 2D-DIGE for biomarker discovery.

Main Methods:

  • Targeted depletion of high-abundance proteins using antibody columns.
Keywords:
AntibodiesDIGEImmunodepletionLow abundantPlasmaSerum

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  • Analysis of depleted plasma samples by fluorescence two-dimensional difference gel electrophoresis (2D-DIGE).
  • Combining immunodepletion with 2D-DIGE for robust proteomic analysis.

    • Main Results:

    • Immunodepletion successfully broadens the coverage of the biofluid proteome.
    • Enhanced detection and quantification of medium-low-abundance proteins are achieved.
    • Facilitates the identification of disease-specific protein biomarkers.

    Conclusions:

    • Upstream immunodepletion is effective for removing high-abundance proteins from plasma.
    • The combination of immunodepletion and 2D-DIGE is a powerful strategy for biomarker discovery.
    • This approach enhances the identification of proteins associated with disease phenotypes.