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Overcoming biofluid protein complexity during targeted mass spectrometry detection and quantification of protein biomarkers by MRM cubed (MRM3)

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Abstract

Targeted mass spectrometry in the so-called multiple reaction monitoring mode (MRM) is certainly a promising way for the precise, accurate, and multiplexed measurement of proteins and their genetic or posttranslationally modified isoforms. MRM carried out on a low-resolution triple quadrupole instrument faces a lack of specificity when addressing the quantification of weakly concentrated proteins. In this case, extensive sample fractionation or immunoenrichment alleviates signal contamination by interferences, but in turn decreases assay performance and throughput. Recently, MRM3 was introduced as an alternative to MRM to improve the limit of quantification of weakly concentrated protein biomarkers. In the present work, we compare MRM and MRM3 modes for the detection of biomarkers in plasma and urine. Calibration curves drawn with MRM and MRM3 showed a similar range of linearity (R 2 > 0.99 for both methods) with protein concentrations above 1 μg/mL in plasma and a few nanogram per milliliter in urine. In contrast, optimized MRM3 methods improve the limits of quantification by a factor of 2 to 4 depending on the targeted peptide. This gain arises from the additional MS3 fragmentation step, which significantly removes or decreases interfering signals within the targeted transition channels.

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References

  1. Kim S-M et al (2011) Identification of peripheral inflammatory markers between normal control and Alzheimer's disease. BMC Neurol 11:51

    Article  CAS  Google Scholar 

  2. Rifai N, Gillette MA, Carr SA (2006) Protein biomarker discovery and validation: the long and uncertain path to clinical utility. Nat Biotechnol 24:971–983

    Article  CAS  Google Scholar 

  3. Zhao Y-N, Dai X-R, Liu J-J, Li X-H, Yang J-J, Sun H, Wu P, Shen J, Lu J-P, Xie H-T, Liu X-Q (2012) An indirect sandwich ELISA for the determination of agkisacutacin in human serum: application to pharmacokinetic study in Chinese healthy volunteers. J Pharm Biomed Anal 70:396–400

    Article  CAS  Google Scholar 

  4. Barton C, Kay RG, Gentzer W, Vitzthum F, Pleasance S (2010) Development of high-throughput chemical extraction techniques and quantitative HPLC-MS/MS (SRM) assays for clinically relevant plasma proteins. J Proteome Res 9:333–340

    Article  CAS  Google Scholar 

  5. Addona TA, Abbatiello SE, Schilling B et al (2009) Multi-site assessment of the precision and reproducibility of multiple reaction monitoring-based measurements of proteins in plasma. Nat Biotechnol 27:633–641

    Article  CAS  Google Scholar 

  6. Domanski D, Percy AJ, Yang J, Chambers AG, Hill JS, Freue GVC, Borchers CH (2012) MRM-based multiplexed quantitation of 67 putative cardiovascular disease biomarkers in human plasma. Proteomics 12:1222–1243

    Article  CAS  Google Scholar 

  7. Parker CE, Domanski D, Percy AJ, Chambers AG, Camenzind AG, Smith DS, Borchers CH (2012) Mass spectrometry in high-throughput clinical biomarker assays: multiple reaction monitoring. Top Curr Chem. doi:10.1007/128_2012_353

    Google Scholar 

  8. Chen Y-T, Chen H-W, Domanski D, Smith DS, Liang K-H, Wu C-C, Chen C-L, Chung T, Chen M-C, Chang Y-S, Parker CE, Borchers CH, Yu J-S (2012) Multiplexed quantification of 63 proteins in human urine by multiple reaction monitoring-based mass spectrometry for discovery of potential bladder cancer biomarkers. J Proteomics 75:3529–3545

    Article  CAS  Google Scholar 

  9. Selevsek N, Matondo M, Sanchez Carbayo M, Aebersold R, Domon B (2011) Systematic quantification of peptides/proteins in urine using selected reaction monitoring. Proteomics 11:1135–1147

    Article  CAS  Google Scholar 

  10. Fortin T, Salvador A, Charrier JP, Lenz C, Lacoux X, Morla A, Choquet-Kastylevsky G, Lemoine J (2009) Clinical quantitation of prostate-specific antigen biomarker in the low nanogram/milliliter range by conventional bore liquid chromatography-tandem mass spectrometry (multiple reaction monitoring) coupling and correlation with ELISA tests. Mol Cell Proteomics 8:1006–1015

    Article  CAS  Google Scholar 

  11. Huillet C, Adrait A, Lebert D, Picard G, Trauchessec M, Louwagie M, Dupuis A, Hittinger L, Ghaleh B, Le Corvoisier P, Jaquinod M, Garin J, Bruley C, Brun V (2012) Accurate quantification of cardiovascular biomarkers in serum using Protein Standard Absolute Quantification (PSAQ™) and selected reaction monitoring. Mol Cell Proteomics 11:1–12

    Article  Google Scholar 

  12. Guerrier L, Fortis F, Boschetti E (2012) Solid-phase fractionation strategies applied to proteomics investigations. Methods Mol Biol 818:11–33

    Article  CAS  Google Scholar 

  13. Lau E, Lam MPY, Siu SO, Kong RPW, Chan WL, Zhou Z, Huang J, Lo C, Chu IK (2011) Combinatorial use of offline SCX and online RP-RP liquid chromatography for iTRAQ-based quantitative proteomics applications. Mol Biosyst 7:1399–1408

    Article  CAS  Google Scholar 

  14. Millioni R, Tolin S, Puricelli L, Sbrignadello S, Fadini GP, Tessari P, Arrigoni G (2011) High abundance proteins depletion vs low abundance proteins enrichment: comparison of methods to reduce the plasma proteome complexity. PLoS One 6:e19603

    Article  CAS  Google Scholar 

  15. Whiteaker JR, Zhao L, Anderson L, Paulovich AG (2010) An automated and multiplexed method for high throughput peptide immunoaffinity enrichment and multiple reaction monitoring mass spectrometry-based quantification of protein biomarkers. Mol Cell Proteomics 9:184–196

    Article  CAS  Google Scholar 

  16. Anderson NL, Anderson NG (2002) The human plasma proteome: history, character, and diagnostic prospects. Mol Cell Proteomics 1:845–867

    Article  CAS  Google Scholar 

  17. Núñez O, Gallart-Ayala H, Martins CPB, Lucci P, Busquets R (2013) State-of-the-art in fast liquid chromatography–mass spectrometry for bio-analytical applications. J Chromatogr B Anal Techn Biomed Life Sci 927:3–21

    Article  Google Scholar 

  18. Swartz ME (2005) UPLC TM: an introduction and review. J Liq Chromatogr Relat Technol 28:1253–1263

    Article  CAS  Google Scholar 

  19. Cortés-Francisco N, Flores C, Moyano E, Caixach J (2011) Accurate mass measurements and ultrahigh-resolution: evaluation of different mass spectrometers for daily routine analysis of small molecules in negative electrospray ionization mode. Anal Bioanal Chem 400:3595–3606

    Article  Google Scholar 

  20. Peterson AC, Russell JD, Bailey DJ, Westphall MS, Coon JJ (2012) Parallel reaction monitoring for high resolution and high mass accuracy quantitative, targeted proteomics. Mol Cell Proteomics 11:1475–1488

    Article  Google Scholar 

  21. Gallien S, Duriez E, Crone C, Kellmann M, Moehring T, Domon B (2012) Targeted proteomic quantification on quadrupole-orbitrap mass spectrometer. Mol Cell Proteomics 11:1709–1723

    Article  Google Scholar 

  22. Gallien S, Duriez E, Demeure K, Domon B (2013) Selectivity of LC-MS/MS analysis: implication for proteomics experiments. J Proteomics 81:148–158

    Article  CAS  Google Scholar 

  23. Liu Y, Hüttenhain R, Surinova S, Gillet LCJ, Mouritsen J, Brunner R, Navarro P, Aebersold R (2013) Quantitative measurements of N-linked glycoproteins in human plasma by SWATH-MS. Proteomics 13:1247–1256

    Article  CAS  Google Scholar 

  24. Kolakowski BM, Mester Z (2007) Review of applications of high-field asymmetric waveform ion mobility spectrometry (FAIMS) and differential mobility spectrometry (DMS). Analyst 132:842–864

    Article  CAS  Google Scholar 

  25. Hatsis P, Kapron JT (2008) A review on the application of high-field asymmetric waveform ion mobility spectrometry (FAIMS) in drug discovery. Rapid Commun Mass Spectrom 22:735–738

    Article  CAS  Google Scholar 

  26. Enjalbert Q, Simon R, Salvador A, Antoine R, Redon S, Ayhan MM, Darbour F, Chambert S, Bretonnière Y, Dugourd P, Lemoine J (2011) Photo-SRM: laser-induced dissociation improves detection selectivity of selected reaction monitoring mode. Rapid Commun Mass Spectrom 25:3375–3381

    Article  CAS  Google Scholar 

  27. Enjalbert Q, Girod M, Simon R, Jeudy J, Chirot F, Salvador A, Antoine R, Dugourd P, Lemoine J (2013) Improved detection specificity for plasma proteins by targeting cysteine-containing peptides with photo-SRM. Anal Bioanal Chem 405:2321–2331

    Article  CAS  Google Scholar 

  28. Fortin T, Salvador A, Charrier JP, Lenz C, Bettsworth F, Lacoux X, Choquet-Kastylevsky G, Lemoine J (2009) Multiple reaction monitoring cubed for protein quantification at the low nanogram/milliliter level in nondepleted human serum. Anal Chem 81:9343–9352

    Article  CAS  Google Scholar 

  29. Lakowski TM, Szeitz A, Pak ML, Thomas D, Vhuiyan MI, Kotthaus J, Clement B, Frankel A (2013) MS3 fragmentation patterns of monomethylarginine species and the quantification of all methylarginine species in yeast using MRM3. J Proteomics 80:43–54

    Article  CAS  Google Scholar 

  30. Albert MA, Glynn RJ, Ridker PM (2003) Plasma concentration of C-reactive protein and the calculated Framingham Coronary Heart Disease Risk Score. Circulation 108:161–165

    Article  CAS  Google Scholar 

  31. Frederiksen C, Lomholt AF, Davis GJ, Dowell BL, Blankenstein MA, Christensen IJ, Brünner N, Nielsen HJ (2009) Changes in plasma TIMP-1 levels after resection for primary colorectal cancer. Anticancer Res 29:75–81

    CAS  Google Scholar 

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Authors and Affiliations

  1. UMR 5280 CNRS Université de Lyon1, Institut des Sciences Analytiques, Université de Lyon, 5 Rue de la Doua, 69100, Villeurbanne, France

    Jeremy Jeudy, Arnaud Salvador, Romain Simon, Aurore Jaffuel, Catherine Fonbonne & Jerome Lemoine

  2. Sanofi, 13 Quai Jules Guesde, 94400, Vitry-sur-Seine, France

    Jean-François Léonard, Jean-Charles Gautier & Olivier Pasquier

Authors
  1. Jeremy Jeudy
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  2. Arnaud Salvador
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  3. Romain Simon
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  4. Aurore Jaffuel
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  5. Catherine Fonbonne
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  6. Jean-François Léonard
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  7. Jean-Charles Gautier
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  8. Olivier Pasquier
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  9. Jerome Lemoine
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Correspondence to Jerome Lemoine.

Additional information

Published in the special issue Analytical Science in France with guest editors Christian Rolando and Philippe Garrigues.

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Jeudy, J., Salvador, A., Simon, R. et al. Overcoming biofluid protein complexity during targeted mass spectrometry detection and quantification of protein biomarkers by MRM cubed (MRM3). Anal Bioanal Chem 406, 1193–1200 (2014). https://doi.org/10.1007/s00216-013-7266-6

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  • DOI: https://doi.org/10.1007/s00216-013-7266-6

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