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Related Experiment Video

Updated: May 21, 2026

Comprehensive Workflow of Mass Spectrometry-based Shotgun Proteomics of Tissue Samples
14:51

Comprehensive Workflow of Mass Spectrometry-based Shotgun Proteomics of Tissue Samples

Published on: November 13, 2021

Robust workflow for iTRAQ-based peptide and protein quantification.

Florian Beck1, Julia Maria Burkhart, Joerg Geiger

  • 1Leibniz-Institut für Analytische Wissenschaften, ISAS-e.V., Dortmund, Germany.

Methods in Molecular Biology (Clifton, N.J.)
|June 6, 2012
PubMed
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This study presents a robust workflow for quantitative proteomics using Isobaric Tagging reagents for Relative and Absolute Quantitation (iTRAQ) labeling. The method enables multiplexed analysis of up to eight samples for comparative protein expression profiling.

Area of Science:

  • Proteomics
  • Analytical Chemistry
  • Biochemistry

Background:

  • Quantitative proteomics is crucial for comparing protein expression in biological samples.
  • Stable isotope labeling techniques like iTRAQ are widely used for this purpose.
  • iTRAQ enables multiplexed quantification at the MS/MS level.

Purpose of the Study:

  • To present a robust workflow for iTRAQ quantification of biological samples.
  • To optimize sample preparation, digestion, labeling, and fractionation for iTRAQ analysis.
  • To facilitate comparative proteomic studies, particularly with human platelets.

Main Methods:

  • Developed a comprehensive workflow including sample preparation and optimized tryptic digestion.
  • Utilized Solid Phase Extraction (SPE) for desalting and a 4-plex iTRAQ labeling kit.

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Selected Reaction Monitoring Mass Spectrometry for Absolute Protein Quantification
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Last Updated: May 21, 2026

Comprehensive Workflow of Mass Spectrometry-based Shotgun Proteomics of Tissue Samples
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Comprehensive Workflow of Mass Spectrometry-based Shotgun Proteomics of Tissue Samples

Published on: November 13, 2021

Selected Reaction Monitoring Mass Spectrometry for Absolute Protein Quantification
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Selected Reaction Monitoring Mass Spectrometry for Absolute Protein Quantification

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  • Fractionated peptide mixtures using strong cation exchange chromatography.

    • Main Results:

    • Established a reliable protocol for iTRAQ-based quantitative proteomics.
    • Demonstrated the applicability of the workflow to complex biological samples like human platelets.
    • The workflow allows for multiplexed analysis of up to eight samples.

    Conclusions:

    • The presented workflow provides a robust method for iTRAQ quantification.
    • This approach enhances the ability to perform comparative proteomic studies.
    • The protocol is suitable for analyzing protein expression profiles in various biological contexts.