Quantitative mass spectrometric multiple reaction monitoring assays for major plasma proteins
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Quantitative LC-MS/MS assays were designed for tryptic peptides representing 53 high and medium abundance proteins in human plasma using a multiplexed multiple reaction monitoring (MRM) approach. Of these, 47 produced acceptable quantitative data, demonstrating within-run coefficients of variation (CVs) (n = 10) of 2-22% (78% of assays had CV <10%). A number of peptides gave CVs in the range 2-7% in five experiments (10 replicate runs each) continuously measuring 137 MRMs, demonstrating the precision achievable in complex digests. Depletion of six high abundance proteins by immunosubtraction significantly improved CVs compared with whole plasma, but analytes could be detected in both sample types. Replicate digest and depletion/digest runs yielded correlation coefficients (R(2)) of 0.995 and 0.989, respectively. Absolute analyte specificity for each peptide was demonstrated using MRM-triggered MS/MS scans. Reliable detection of L-selectin (measured at 0.67 microg/ml) indicates that proteins down to the microg/ml level can be quantitated in plasma with minimal sample preparation, yielding a dynamic range of 4.5 orders of magnitude in a single experiment. Peptide MRM measurements in plasma digests thus provide a rapid and specific assay platform for biomarker validation, one that can be extended to lower abundance proteins by enrichment of specific target peptides (stable isotope standards and capture by anti-peptide antibodies (SISCAPA)).
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LC-MS/MS is suitable for highly multiplexed protein quantification and isoform differentiation, including apolipoproteins.
“LC-MS/MS has proven suitable for multiplexed quantification and phenotyping of apolipoproteins.”
From Automated multiplex LC-MS/MS assay for quantifying serum apolipoproteins A-I, B, C-I, C-II, C-III, and E with qualitative apolipoprotein E phenotyping (2016) · cites this paper as [11]
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- Authors
- Anderson, L., & Hunter, C. L.
- Corresponding author
- Anderson, Leigh
- ORCID
- Hunter, Christie L.
- Venue
- Molecular & Cellular Proteomics
- Publisher
- Elsevier BV
- Published
- 2006-04
- Volume
- 5
- Issue
- 4
- Pages
- 573-588
- ISSN
- 1535-9476 (ISSN-L)
- OpenAlex
- W2056467018
- Cited by
- 1,230
- References
- 1
- Citations by year
- 2026: 10 · 2025: 12 · 2024: 19 · 2023: 13 · 2022: 14 · 2021: 15 · 2020: 17 · 2019: 28 · 2018: 41 · 2017: 57 · 2016: 63 · 2015: 70 · 2014: 93 · 2013: 124 · 2012: 139
- FWCI
- 25.04
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Molecular & Cellular Proteomics (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Metabolomics and Mass Spectrometry Studies
- Keywords
- plasma proteins, multiple reaction monitoring, quantitative proteomics, high-abundance protein depletion, L-selectin, tryptic peptides, biomarker validation, peptide enrichment, LC-MS/MS method development
- MeSH terms
- Amino Acid Sequence, Blood Proteins, Chromatography, Liquid, Humans, Molecular Sequence Data, Peptide Mapping, Reference Standards, Sensitivity and Specificity, Reproducibility of Results, Spectrometry, Mass, Electrospray Ionization
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