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Related Experiment Video

Updated: Jun 21, 2026

Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification
10:37

Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification

Published on: November 15, 2017

A technically detailed and pragmatic protocol for quantitative serum proteomics using iTRAQ.

Sarah Tonack1, Mark Aspinall-O'Dea, Rosalind E Jenkins

  • 1Royal Liverpool University Hospital, University of Liverpool, United Kingdom.

Journal of Proteomics
|August 5, 2009
PubMed
Summary

This study presents a refined protocol for quantifying serum proteins using isobaric tags for relative and absolute quantification (iTRAQ) after removing high-abundance proteins. The improved method enhances biomarker discovery for diseases like pancreatic cancer.

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Area of Science:

  • Biochemistry
  • Proteomics
  • Biomarker Discovery

Background:

  • Blood, particularly serum and plasma, is a crucial source of disease biomarkers.
  • Analyzing the complex protein concentration range in serum for biomarkers is challenging.
  • Existing proteomic methods require simplification strategies and advanced quantification techniques.

Purpose of the Study:

  • To provide a detailed protocol for iTRAQ-based quantification of serum proteins.
  • To optimize the iTRAQ protocol for enhanced protein identification and quantification.
  • To apply the protocol to serum samples from pancreatic cancer patients.

Main Methods:

  • Immunodepletion of high-abundance proteins from serum samples.
  • Application of a modified isobaric tags for relative and absolute quantification (iTRAQ) protocol.
  • Proteomic analysis focusing on peptide identification and relative quantification.

Main Results:

  • Identification of 217 proteins (5773 peptides) at a 1% false discovery rate.
  • Identification of 254 proteins with 95% confidence.
  • Relative quantification of 234 serum proteins (95% confidence), including low-abundance markers.

Conclusions:

  • The modified iTRAQ protocol successfully quantifies a higher number of serum proteins.
  • This method is effective for identifying potential biomarkers in diseases like pancreatic cancer.
  • The protocol is adaptable for serum proteomic analysis across various disease types.