Abstract
LC-MS/MS, particularly when linked with immunoaffinity enrichment, has emerged as a highly capable bioanalytical technique for the quantitative measurement of protein biomarkers and therapeutic proteins, thus impacting translational pharmacology. A key advantage of a protein LC-MS/MS assay over other bioanalytical techniques is the high measurement specificity that can be achieved. Immunoaffinity enrichment techniques using anti-protein or anti-peptide antibodies, or both in a sequential manner, extend LC-MS/MS assay sensitivity for protein biomarkers into the pg/mL range. Assay translation between species can be facilitated by selecting proteotypic peptides that are conserved in the same protein across species, if available, to allow the same MS detection method, the same SIL standard peptide and the same anti-peptide antibody can be used. Practical challenges to routine implementation in clinical assays are being overcome by the use of standardized workflows, liquid handling robotics, and robust LC-MS/MS configurations.
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Abbreviations
- Translational pharmacology :
-
Investigations of drug effects on pathways and disease to establish a mechanistic link between in vitro or ex vivo to in vivo systems as well as within and between species. For example, to identify in vivo pharmacology and biomarkers in preclinical species that can also be measured in humans
- Proteotypic peptide :
-
A peptide, enzymatically released from a protein as part of an LC-MS/MS assay, which serves to unambiguously identify that protein
- Selected reaction monitoring :
-
Selected reaction monitoring (SRM) is a tandem mass spectrometry technique in which an ion of specified mass-to-charge ratio is selected in the first mass spectrometry stage. One or several product ions of a specified mass-to-charge ratio resulting from fragmentation of the precursor are detected in the second mass spectrometry stage
- Sequential protein and peptide immunoaffinity :
-
Dual immunoaffinity sample preparation technique used for measuring low abundance protein biomarkers in biological matrices by mass spectrometry. Protein immunoaffinity enrichment of the protein biomarker using an anti-protein capture reagent is followed by digestion and immunoaffinity enrichment of one or several enzymatically released peptides using anti-peptide antibodies
References
Rifai N, Gillette MA, Carr SA (2006) Protein biomarker discovery and validation: the long and uncertain path to clinical utility. Nat Biotechnol 24(8):971–983
Mallick P, Kuster B (2010) Proteomics: a pragmatic perspective. Nat Biotechnol 28(7):695–709
Coombs KM (2011) Quantitative proteomics of complex mixtures. Expert Rev Proteomics 8(5):659–677
Egertson JD, Kuehn A, Merrihew GE et al (2013) Multiplexed MS/MS for improved data-independent acquisition. Nat Methods 10(8):744–746
Fernandez Ocaña M, Neubert H, Przyborowska A et al (2004) BSE control: detection of gelatine-derived peptides in animal feed by mass spectrometry. Analyst 129(2):111–115
Neubert H, Grace C, Rumpel K, James I (2008) Assessing immunogenicity in the presence of excess protein therapeutic using immunoprecipitation and quantitative mass spectrometry. Anal Chem 80(18):6907–6914
Neubert H, James I (2009) Online capillary weak cation exchange enrichment hyphenated to nanospray mass spectrometry for quantitation of a basic pegvisomant derived peptide. J Chromatogr A 1216(33):6151–6154
Anderson NL, Anderson NG (2002) The human plasma proteome: history, character, and diagnostic prospects. Mol Cell Proteomics 1(11):845–867
Jian W, Edom RW, Wang D, Weng N, Zhang S (2013) Relative quantitation of glycoisoforms of intact apolipoprotein C3 in human plasma by liquid chromatography-high-resolution mass spectrometry. Anal Chem 85(5):2867–2874
Ackermann BL, Berna MJ (2007) Coupling immunoaffinity techniques with MS for quantitative analysis of low-abundance protein biomarkers. Expert Rev Proteomics 4(2):175–186
Neubert H, Muirhead D, Kabir M, Grace C, Cleton A, Arends R (2013) Sequential protein and peptide immunoaffinity capture for mass spectrometry-based quantification of total human beta-nerve growth factor. Anal Chem 85(3):1719–1726
Palandra J, Finelli A, Zhu M, Masferrer J, Neubert H (2013) Highly specific and sensitive measurements of human and monkey interleukin 21 using sequential protein and tryptic peptide immunoaffinity LC-MS/MS. Anal Chem 85(11):5522–5529
Fernández Ocaña M, Neubert H (2010) An immunoaffinity liquid chromatography-tandem mass spectrometry assay for the quantitation of matrix metalloproteinase 9 in mouse serum. Anal Biochem 399(2):202–210
Schultz GA, Mccardle K, Neubert H (2016) Large-scale implementation of sequential protein and peptide immunoaffinity enrichment LC/nanoLC–MS/MS for human β-nerve growth factor. Bioanalysis 8(8):753–764
Palandra J, Quazi A, Fitz L, Rong H, Morris C, Neubert H (2016) Quantitative measurements of GDF-8 using Immunoaffinity LC-MS/MS. Proteomics Clin Appl 10(5):597–604. doi:10.1002/prca.201500112n/a-n/a
Anderson NL, Anderson NG, Haines LR, Hardie DB, Olafson RW, Pearson TW (2004) Mass spectrometric quantitation of peptides and proteins using Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA). J Proteome Res 3(2):235–244
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(1):184–196
Neubert H, Gale J, Muirhead D (2010) Online high-flow peptide immunoaffinity enrichment and nanoflow LC-MS/MS: assay development for total salivary pepsin/pepsinogen. Clin Chem 56(9):1413–1423
Shah B, Reid J, Kuzyk M, Parker C, Borchers C (2013) Developing an iMALDI Method. Methods Mol Biol 1023:97–120
Fan Y-Y, Neubert H (2016) Quantitative Analysis of Human Neonatal Fc Receptor (FcRn) tissue expression in transgenic mice by online peptide immuno-affinity LC-HRMS. Anal Chem. doi:10.1021/acs.analchem.5b03900
Wang H, Bennett P (2013) Performance assessment of microflow LC combined with high-resolution MS in bioanalysis. Bioanalysis 5(10):1249–1267
Lee AYH, Chappell DL, Bak MJ et al (2016) Multiplexed quantification of proglucagon-derived peptides by immunoaffinity enrichment and tandem mass spectrometry after a meal tolerance test. Clin Chem 62(1):227–235
Picotti P, Aebersold R (2012) Selected reaction monitoring-based proteomics: workflows, potential, pitfalls and future directions. Nat. Meth. 9(6):555–566
Sherman J, Mckay MJ, Ashman K, Molloy MP (2009) How specific is my SRM? The issue of precursor and product ion redundancy. Proteomics 9(5):1120–1123
Hoofnagle AN, Whiteaker JR, Carr SA et al (2016) Recommendations for the generation, quantification, storage, and handling of peptides used for mass spectrometry-based assays. Clin Chem 62(1):48–69
Pope ME, Soste MV, Eyford BA, Anderson NL, Pearson TW (2009) Anti-peptide antibody screening: selection of high affinity monoclonal reagents by a refined surface plasmon resonance technique. J Immunol Methods 341(1–2):86–96
Ackermann BL (2012) Understanding the role of immunoaffinity-based mass spectrometry methods for clinical applications. Clin Chem 58(12):1620–1622
Lee J, Devanarayan V, Barrett Y et al (2006) Fit-for-purpose method development and validation for successful biomarker measurement. Pharm Res 23(2):312–328
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Neubert, H. (2016). Quantification of Protein Biomarkers Using Liquid Chromatography Tandem Mass Spectrometry. In: Weiner, R., Kelley, M. (eds) Translating Molecular Biomarkers into Clinical Assays . AAPS Advances in the Pharmaceutical Sciences Series, vol 21. Springer, Cham. https://doi.org/10.1007/978-3-319-40793-7_9
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DOI: https://doi.org/10.1007/978-3-319-40793-7_9
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