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Considerations for Soluble Protein Biomarker Blood Sample Matrix Selection

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Abstract

Blood-based soluble protein biomarkers provide invaluable clinical information about patients and are used as diagnostic, prognostic, and pharmacodynamic markers. The most commonly used blood sample matrices are serum and different types of plasma. In drug development research, the impact of sample matrix selection on successful protein biomarker quantification is sometimes overlooked. The sample matrix for a specific analyte is often chosen based on prior experience or literature searches, without good understanding of the possible effects on analyte quantification. Using a data set of 32 different soluble protein markers measured in matched serum and plasma samples, we examined the differences between serum and plasma and discussed how platelet or immune cell activation can change the quantified concentration of the analyte. We have also reviewed the effect of anticoagulant on analyte quantification. Finally, we provide specific recommendations for biomarker sample matrix selection and propose a systematic and data-driven approach for sample matrix selection. This review is intended to raise awareness of the impact and considerations of sample matrix selection on biomarker quantification.

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References

  1. Ladenson JH, Tsai L-MB, Michael JM, Kessler G, Joist JH. Serum versus heparinized plasma for eighteen common chemistry tests: is serum the appropriate specimen? Am J Clin Pathol. 1974;62(4):545–52.

    CAS  PubMed  Google Scholar 

  2. Miles RR, Roberts RF, Putnam AR, Roberts WL. Comparison of serum and heparinized plasma samples for measurement of chemistry analytes. Clin Chem. 2004;50(9):1704–6.

    CAS  PubMed  Google Scholar 

  3. Dupin M, Fortin T, Larue-Triolet A, Surault I, Beaulieu C, Gouel-Chéron A, et al. Impact of serum and plasma matrices on the titration of human inflammatory biomarkers using analytically validated SRM assays. J Proteome Res. 2016;15(8):2366–78.

    CAS  PubMed  Google Scholar 

  4. Jung K, Laube C, Lein M, Lichtinghagen R, Tschesche H, Schnorr D, et al. Kind of sample as preanalytical determinant of matrix metalloproteinases 2 and 9 and tissue inhibitor of metalloproteinase 2 in blood. Clin Chem. 1998;44(5):1060–2.

    CAS  PubMed  Google Scholar 

  5. Scholman RC, Giovannone B, Hiddingh S, Meerding JM, Malvar Fernandez B, van Dijk MEA, et al. Effect of anticoagulants on 162 circulating immune related proteins in healthy subjects. Cytokine. 2018;106:114–24.

    CAS  PubMed  Google Scholar 

  6. Biancotto A, Feng X, Langweiler M, Young NS, McCoy JP. Effect of anticoagulants on multiplexed measurement of cytokine/chemokines in healthy subjects. Cytokine. 2012;60(2):438–46.

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Brondum L, Sorensen BS, Eriksen JG, Mortensen LS, Lonbro S, Overgaard J, et al. An evaluation of multiplex bead-based analysis of cytokines and soluble proteins in archived lithium heparin plasma, EDTA plasma and serum samples. Scand J Clin Lab Invest. 2016;76(8):601–11.

    PubMed  Google Scholar 

  8. Tvedt TH, Rye KP, Reikvam H, Brenner AK, Bruserud O. The importance of sample collection when using single cytokine levels and systemic cytokine profiles as biomarkers--a comparative study of serum versus plasma samples. J Immunol Methods. 2015;418:19–28.

    CAS  PubMed  Google Scholar 

  9. Keustermans GC, Hoeks SB, Meerding JM, Prakken BJ, de Jager W. Cytokine assays: an assessment of the preparation and treatment of blood and tissue samples. Methods. 2013;61(1):10–7.

    CAS  PubMed  Google Scholar 

  10. Thavasu PW, Longhurst S, Joel SP, Slevin ML, Balkwill FR. Measuring cytokine levels in blood. Importance of anticoagulants, processing, and storage conditions. J Immunol Methods. 1992;153(1–2):115–24.

    CAS  PubMed  Google Scholar 

  11. Hennø LT, Storjord E, Christiansen D, Bergseth G, Ludviksen JK, Fure H, et al. Effect of the anticoagulant, storage time and temperature of blood samples on the concentrations of 27 multiplex assayed cytokines – consequences for defining reference values in healthy humans. Cytokine. 2017;97:86–95.

    PubMed  Google Scholar 

  12. O’Neal WK, Anderson W, Basta PV, Carretta EE, Doerschuk CM, Barr RG, et al. of serum, EDTA plasma and P100 plasma for luminex-based biomarker multiplex assays in patients with chronic obstructive pulmonary disease in the SPIROMICS study. J Transl Med. 2014;12(1):9.

    PubMed  PubMed Central  Google Scholar 

  13. de Jager W, Bourcier K, Rijkers GT, Prakken BJ, Seyfert-Margolis V. Prerequisites for cytokine measurements in clinical trials with multiplex immunoassays. BMC Immunol. 2009;10(1):52.

    PubMed  PubMed Central  Google Scholar 

  14. Lan J, Nunez Galindo A, Doecke J, Fowler C, Martins RN, Rainey-Smith SR, et al. Systematic evaluation of the use of human plasma and serum for mass-spectrometry-based shotgun proteomics. J Proteome Res. 2018;17(4):1426–35.

    CAS  PubMed  Google Scholar 

  15. Ilies M, Iuga CA, Loghin F, Dhople VM, Thiele T, Völker U, et al. Data on the impact of the blood sample collection methods on blood protein profiling studies. Data Brief. 2017;14:313–9.

    PubMed  PubMed Central  Google Scholar 

  16. Organization WH. Use of anticoagulants in diagnostic laboratory investigations. 2nd ed. Geneva: World Health Organization; 2002.

    Google Scholar 

  17. Nossel HL. Differential consumption of coagulation factors resulting from activation of the extrinsic (tissue thromboplastin) or the intrinsic (foreign surface contact) pathways. Blood. 1967;29(3):331–40.

    CAS  PubMed  Google Scholar 

  18. Carey RN, Jani C, Johnson C, Pearce J, Hui-Ng P, Lacson E. Chemistry testing on plasma versus serum samples in dialysis patients: clinical and quality improvement implications. Clin J Am Soc Nephrol. 2016;11(9):1675–9.

    PubMed  PubMed Central  Google Scholar 

  19. Smith SA, Travers RJ, Morrissey JH. How it all starts: initiation of the clotting cascade. Crit Rev Biochem Mol Biol. 2015;50(4):326–36.

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Machlus KR, Johnson KE, Kulenthirarajan R, Forward JA, Tippy MD, Soussou TS, et al. CCL5 derived from platelets increases megakaryocyte proplatelet formation. Blood. 2016;127(7):921–6.

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Power CA, Clemetson JM, Clemetson KJ, Wells TNC. Chemokine and chemokine receptor mRNA expression in human platelets. Cytokine. 1995;7(6):479–82.

    CAS  PubMed  Google Scholar 

  22. Zhao X, Delgado L, Weiner R, Laterza OF. Influence of pre-analytical factors on thymus- and activation-regulated chemokine quantitation in plasma. J Circ Biomark. 2015;4:10.

    PubMed  PubMed Central  Google Scholar 

  23. Assoian RK, Sporn MB. Type beta transforming growth factor in human platelets: release during platelet degranulation and action on vascular smooth muscle cells. J Cell Biol. 1986;102(4):1217–23.

    CAS  PubMed  Google Scholar 

  24. Otterdal K, Smith C, Øie E, Pedersen TM, Yndestad A, Stang E, et al. Platelet-derived LIGHT induces inflammatory responses in endothelial cells and monocytes. Blood. 2006;108(3):928–35.

    CAS  PubMed  Google Scholar 

  25. Severin IC, Gaudry JP, Johnson Z, Kungl A, Jansma A, Gesslbauer B, et al. Characterization of the chemokine CXCL11-heparin interaction suggests two different affinities for glycosaminoglycans. J Biol Chem. 2010;285(23):17713–24.

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Mei J, Liu Y, Dai N, Favara M, Greene T, Jeyaseelan S, et al. CXCL5 regulates chemokine scavenging and pulmonary host defense to bacterial infection. Immunity. 2010;33(1):106–17.

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Salanga CL, Dyer DP, Kiselar JG, Gupta S, Chance MR, Handel TM. Multiple glycosaminoglycan-binding epitopes of monocyte chemoattractant protein-3/CCL7 enable it to function as a non-oligomerizing chemokine. J Biol Chem. 2014;289(21):14896–912.

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Seo Y, Schenauer MR, Leary JA. Biologically relevant metal-cation binding induces conformational changes in heparin oligosaccharides as measured by ion mobility mass spectrometry. Int J Mass Spectrom. 2011;303(2–3):191–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Visse R, Nagase H. Matrix metalloproteinases and tissue inhibitors of metalloproteinases. Circ Res. 2003;92(8):827–39.

    CAS  PubMed  Google Scholar 

  30. Tallant C, Marrero A, Gomis-Rüth FX. Matrix metalloproteinases: fold and function of their catalytic domains. Biochim Biophys Acta. 2010;1803(1):20–8.

    CAS  PubMed  Google Scholar 

  31. Tezvergil-Mutluay A, Agee KA, Hoshika T, Carrilho M, Breschi L, Tjäderhane L, et al. The requirement of zinc and calcium ions for functional MMP activity in demineralized dentin matrices. Dent Mater. 2010;26(11):1059–67.

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Paula K, Anne T-M, Taina T-H. The sample type used affects the levels of gelatinases (MMP-2 and -9) and their inhibitors (TIMP-1 and -2) in circulating blood of healthy controls and breast cancer patients. Biomark Insights. 2007;2:117–27.

    PubMed  PubMed Central  Google Scholar 

  33. John M, Jung K. Pre-analytical conditions for the assessment of circulating MMP-9 and TIMP-1: consideration of pitfalls. Eur Respir J. 2005;26(2):364–5.

    CAS  PubMed  Google Scholar 

  34. Gerlach RF, Uzuelli JA, Souza-Tarla CD, Tanus-Santos JE. Effect of anticoagulants on the determination of plasma matrix metalloproteinase (MMP)-2 and MMP-9 activities. Anal Biochem. 2005;344(1):147–9.

    CAS  PubMed  Google Scholar 

  35. Fischer JE, Janousek M, Fischer M, Seifarth FG, Blau N, Fanconi S. Effect of collection and preprocessing methods on neutrophil elastase plasma concentrations. Clin Biochem. 1998;31(3):131–6.

    CAS  PubMed  Google Scholar 

  36. Beatty K, Bieth J, Travis J. Kinetics of association of serine proteinases with native and oxidized alpha-1-proteinase inhibitor and alpha-1-antichymotrypsin. J Biol Chem. 1980;255(9):3931–4.

    CAS  PubMed  Google Scholar 

  37. Frommherz KJ, Faller B, Bieth JG. Heparin strongly decreases the rate of inhibition of neutrophil elastase by alpha 1-proteinase inhibitor. J Biol Chem. 1991;266(23):15356–62.

    CAS  PubMed  Google Scholar 

  38. Ellyard JI, Simson L, Bezos A, Johnston K, Freeman C, Parish CR. Eotaxin selectively binds heparin: an interaction that protects eotaxin from proteolysis and potentiates chemotactic activity in vivo. J Biol Chem. 2007;282(20):15238–47.

    CAS  PubMed  Google Scholar 

  39. Cedrone E, Neun BW, Rodriguez J, Vermilya A, Clogston JD, McNeil SE, et al. Anticoagulants influence the performance of in vitro assays intended for characterization of nanotechnology-based formulations. Molecules. 2017;23(1).

  40. Ray CA. Biomarker accuracy: exploring the truth. Bioanalysis. 2014;6(3):269–71.

    CAS  PubMed  Google Scholar 

  41. Smyth SS, McEver RP, Weyrich AS, Morrell CN, Hoffman MR, Arepally GM, et al. Platelet functions beyond hemostasis. J Thromb Haemost. 2009;7(11):1759–66.

    CAS  PubMed  Google Scholar 

  42. Burkhart JM, Gambaryan S, Watson SP, Jurk K, Walter U, Sickmann A, et al. What can proteomics tell us about platelets? Circ Res. 2014;114(7):1204–19.

    CAS  PubMed  Google Scholar 

  43. Fong KP, Barry C, Tran AN, Traxler EA, Wannemacher KM, Tang H-Y, et al. Deciphering the human platelet sheddome. Blood. 2011;117(1):e15–26.

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Senzel L, Gnatenko DV, Bahou WF. The platelet proteome. Curr Opin Hematol. 2009;16(5):329–33.

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Coppinger JA, Cagney G, Toomey S, Kislinger T, Belton O, McRedmond JP, et al. Characterization of the proteins released from activated platelets leads to localization of novel platelet proteins in human atherosclerotic lesions. Blood. 2004;103(6):2096–104.

    CAS  PubMed  Google Scholar 

  46. Kong F-MS, Zhao L, Wang L, Chen Y, Hu J, Fu X, et al. Ensuring sample quality for blood biomarker studies in clinical trials: a multicenter international study for plasma and serum sample preparation. Transl Lung Cancer Res. 2017;6(6):625–34.

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Jonnalagadda D, Izu LT, Whiteheart SW. Platelet secretion is kinetically heterogeneous in an agonist-responsive manner. Blood. 2012;120(26):5209–16.

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Ding D, Liu X, Duan J, Guo S-W. Platelets are an unindicted culprit in the development of endometriosis: clinical and experimental evidence. Hum Reprod. 2015;30(4):812–32.

    CAS  PubMed  Google Scholar 

  49. Gresele P, Fuster V, Lopez JA, Page CP, Vermylen J, editors. Platelets in hematologic and cardiovascular disorders. 1st ed: Cambridge University Press; 2008.

  50. Peterson JE, Zurakowski D, Italiano JE, Michel LV, Fox L, Klement GL, et al. Normal ranges of angiogenesis regulatory proteins in human platelets. Am J Hematol. 2010;85(7):487–93.

    CAS  PubMed  Google Scholar 

  51. Halldórsdóttir AM, Stoker J, Porche-Sorbet R, Eby CS. Soluble CD40 ligand measurement inaccuracies attributable to specimen type, processing time, and ELISA method. Clin Chem. 2005;51(6):1054–7.

    PubMed  Google Scholar 

  52. Zhao X, Delgado L, Weiner R, Laterza OF. An ultra-sensitive clinical biomarker assay: quantitation of thymus and activation-regulated chemokine in human plasma. Bioanalysis. 2014;6(8):1069–80.

    CAS  PubMed  Google Scholar 

  53. Wakefield LM, Letterio JJ, Chen T, Danielpour D, Allison RS, Pai LH, et al. Transforming growth factor-beta1 circulates in normal human plasma and is unchanged in advanced metastatic breast cancer. Clin Cancer Res. 1995;1(1):129–36.

    CAS  PubMed  Google Scholar 

  54. O’Brien PJ, Ramanathan R, Yingling JM, Baselga J, Rothenberg ML, Carducci M, et al. Analysis and variability of TGFbeta measurements in cancer patients with skeletal metastases. Biologics. 2008;2(3):563–9.

    PubMed  PubMed Central  Google Scholar 

  55. Swystun LL, Liaw PC. The role of leukocytes in thrombosis. Blood. 2016;128(6):753–62.

    CAS  PubMed  Google Scholar 

  56. von Brühl M-L, Stark K, Steinhart A, Chandraratne S, Konrad I, Lorenz M, et al. Monocytes, neutrophils, and platelets cooperate to initiate and propagate venous thrombosis in mice in vivo. J Exp Med. 2012;209(4):819.

    Google Scholar 

  57. Kambas K, Mitroulis I, Ritis K. The emerging role of neutrophils in thrombosis-the journey of TF through NETs. Front Immunol. 2012;3:385.

    PubMed  PubMed Central  Google Scholar 

  58. Weisel JW, Litvinov RI. Red blood cells: the forgotten player in hemostasis and thrombosis. J Thromb Haemost. 2019;17(2):271–82.

    CAS  PubMed  PubMed Central  Google Scholar 

  59. Smiley ST, King JA, Hancock WW. Fibrinogen stimulates macrophage chemokine secretion through toll-like receptor 4. J Immunol. 2001;167(5):2887.

    CAS  PubMed  Google Scholar 

  60. Parkitny L, McAuley JH, Kelly PJ, Di Pietro F, Cameron B, Moseley GL. Multiplex cytokine concentration measurement: how much do the medium and handling matter? Mediat Inflamm. 2013;2013:13.

    Google Scholar 

  61. Ryu JK, Petersen MA, Murray SG, Baeten KM, Meyer-Franke A, Chan JP, et al. Blood coagulation protein fibrinogen promotes autoimmunity and demyelination via chemokine release and antigen presentation. Nat Commun. 2015;6:8164.

    PubMed  PubMed Central  Google Scholar 

  62. Koller DY. Sampling methods. Am J Respir Crit Care Med. 2000;162(supplement_1):S31–S3.

    CAS  PubMed  Google Scholar 

  63. Jonsson A, Hjalmarsson C, Falk P, Ivarsson M-L. Levels of matrix metalloproteinases differ in plasma and serum - aspects regarding analysis of biological markers in cancer. Br J Cancer. 2016;115(6):703–6.

    CAS  PubMed  PubMed Central  Google Scholar 

  64. Freitas M, Porto G, Lima JLFC, Fernandes E. Isolation and activation of human neutrophils in vitro. The importance of the anticoagulant used during blood collection. Clin Biochem. 2008;41(7):570–5.

    CAS  PubMed  Google Scholar 

  65. Lee JW, Devanarayan V, Barrett YC, Weiner R, Allinson J, Fountain S, et al. Fit-for-purpose method development and validation for successful biomarker measurement. Pharm Res. 2006;23(2):312–28.

    CAS  PubMed  Google Scholar 

  66. Booth B, Arnold ME, DeSilva B, Amaravadi L, Dudal S, Fluhler E, et al. Workshop report: Crystal City V--quantitative bioanalytical method validation and implementation: the 2013 revised FDA guidance. AAPS J. 2014;17(2):277–88.

    PubMed  PubMed Central  Google Scholar 

  67. Arnold ME, Booth B, King L, Ray C. Workshop report: Crystal City VI—bioanalytical method validation for biomarkers. AAPS J. 2016;18(6):1366–72.

    CAS  PubMed  Google Scholar 

  68. Piccoli SP, Garofolo F. Biomarker assay validation. Bioanalysis. 2018;10(12):889–91.

    CAS  PubMed  Google Scholar 

  69. Bowen RAR, Remaley AT. Interferences from blood collection tube components on clinical chemistry assays. Biochemia Med. 2014;24(1):31–44.

    CAS  Google Scholar 

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Acknowledgments

We would like to thank Jennifer Lawson for all the work she did in gathering the data used in Tables I and II and supplementary material. Additionally, we would like to thank Ramu Thiruvamoor and Jason Y. Park who helped in the initial process of gathering data and case studies for this paper.

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Authors and Affiliations

  1. Ionis Pharmaceuticals, 2855 Gazelle Rd., Carlsbad, California, 92010, USA

    Joel A. Mathews

  2. Regeneron, Tarrytown, New York, USA

    Yan G. Ni, Daoyu Duan & Sara Hamon

  3. Novartis, Cambridge, Massachusetts, USA

    Connie Wang

  4. Bristol-Myers Squibb, Princeton, New Jersey, USA

    Jon E. Peterson

  5. Zoetis, Kalamazoo, Michigan, USA

    Chad Ray

  6. Merck & Co., Kenilworth, New Jersey, USA

    Xuemei Zhao

  7. Immunologix Laboratories, Tampa, Florida, USA

    John Allinson

  8. Genentech, South San Francisco, California, USA

    Martha Hokom

  9. R&D Systems, Minneapolis, Minnesota, USA

    Greta Wegner

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  1. Joel A. Mathews
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  2. Yan G. Ni
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Correspondence to Joel A. Mathews.

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Mathews, J.A., Ni, Y.G., Wang, C. et al. Considerations for Soluble Protein Biomarker Blood Sample Matrix Selection. AAPS J 22, 38 (2020). https://doi.org/10.1208/s12248-020-0412-0

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