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Updated: Jun 17, 2026
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Deep Proteome Profiling by Isobaric Labeling, Extensive Liquid Chromatography, Mass Spectrometry, and Software-assisted Quantification
Published on: November 15, 2017
iTRAQ-based quantitative analysis of protein mixtures with large fold change and dynamic range
Juan Casado-Vela1, María José Martínez-Esteso, Eva Rodriguez
1Proteomics Platform, CIC bioGUNE, CIBERehd, ProteoRed, Technology Park of Bizkaia, Derio, Spain. jcasado@cicbiogune.es
Proteomics
|December 24, 2009
Summary
New protein mixtures enable accurate relative protein quantitation using isobaric tag for relative and absolute quantitation (iTRAQ). These samples address challenges in high-dynamic-range proteomic analysis for biomarker discovery.
Area of Science:
- Proteomics
- Biomarker Discovery
- Mass Spectrometry
Background:
- Accurate quantitation of protein abundance changes is crucial for understanding biological systems and identifying novel biomarkers.
- Current HPLC-MS/MS methods face limitations in quantifying protein mixtures due to wide expression ranges and high dynamic concentration ranges.
- Developing reliable quantitation algorithms and standardized samples is essential for advancing proteomic analysis.
Purpose of the Study:
- To introduce two novel protein mixture samples (4-protmix and 8-protmix) designed for relative protein quantitation using isobaric tag for relative and absolute quantitation (iTRAQ).
- To evaluate the suitability of these samples for assessing the performance of iTRAQ-based quantitative proteomic workflows.
Main Methods:
- Development of two distinct protein mixtures (4-protmix and 8-protmix) with varying concentration ranges and relative abundances.
- Analysis of iTRAQ-labeled samples using LTQ Orbitrap mass spectrometry with higher energy collision-induced dissociation (HCD) fragmentation.
- Quantification of protein abundance changes using Proteome Discoverer software.
Main Results:
- The 4-protmix sample enabled measurement of relative protein changes up to 24-fold.
- The 8-protmix sample facilitated quantitation of relative protein changes across two orders of magnitude in concentration and tenfold differences in abundance.
- Demonstrated the utility of the developed samples for testing iTRAQ quantitative proteomic workflows.
Conclusions:
- The presented 4-protmix and 8-protmix samples are valuable tools for validating and optimizing iTRAQ-based relative protein quantitation strategies.
- These standardized mixtures can enhance the reliability and reproducibility of proteomic analyses in diverse laboratory settings.
- Facilitates more accurate biomarker discovery and a deeper understanding of biological system alterations.
