Abstract
To standardize C-reactive protein (CRP) assays, the National Metrology Institute of Japan (NMIJ) has developed a C-reactive protein solution certified reference material, CRM 6201-b, which is intended for use as a primary reference material to enable the SI-traceable measurement of CRP. This study describes the development process of CRM 6201-b. As a candidate material of the CRM, recombinant human CRP solution was selected because of its higher purity and homogeneity than the purified material from human serum. Gel filtration chromatography was used to examine the homogeneity and stability of the present CRM. The total protein concentration of CRP in the present CRM was determined by amino acid analysis coupled to isotope-dilution mass spectrometry (IDMS-AAA). To improve the accuracy of IDMS-AAA, we optimized the hydrolysis process by examining the effect of parameters such as the volume of protein samples taken for hydrolysis, the procedure of sample preparation prior to the hydrolysis, hydrolysis temperature, and hydrolysis time. Under optimized conditions, we conducted two independent approaches in which the following independent hydrolysis and liquid chromatography-isotope dilution mass spectrometry (LC-IDMS) were combined: one was vapor-phase acid hydrolysis (130 °C, 24 h) and hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-MS) method, and the other was microwave-assisted liquid-phase acid hydrolysis (150 °C, 3 h) and pre-column derivatization liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. The quantitative values of the two different amino acid analyses were in agreement within their uncertainties. The certified value was the weighted mean of the results of the two methods. Uncertainties from the value-assignment method, between-method variance, homogeneity, long-term stability, and short-term stability were taken into account in evaluating the uncertainty for a certified value. The certified value and the expanded uncertainty (k = 2) of CRM 6201-b are (40.0 ± 1.6) μmol kg–1.
References
Roux KH, Kilpatrick JM, Volanakis JE, Kearney JF (1983) Localization of the phosphocholine-binding sites on C-reactive protein by immunoelectron microscopy. J Immunol 131:2411–2415
Shrive AK, Cheetham GMT, Holden D, Myles DAA, Turnell WG, Volanakis JE, Pepys MB, Bloomer AC, Greenhough TJ (1996) Three-dimensional structure of human C-reactive protein. Nat Struct Biol 3:346–354
Pepys MB, Hirschfield GM (2003) C-reactive protein: a critical update. J Clin Invest 111:1805–1812
Rifai N, Ridker PM (2001) High-sensitivity C-reactive protein: a novel and promising marker of coronary heart disease. Clin Chem 47:403–411
Danesh J, Whincup P, Walker M, Lennon L, Thomson A, Appleby P, Gallimore JR, Pepys MB (2000) Low grade inflammation and coronary heart disease: prospective study and updated meta-analyses. BMJ 321:199–204
Retterstol L, Eikvar L, Bohn M, Bakken A, Erikssen J, Berg K (2002) C-reactive protein predicts death in patients with previous premature myocardial infarction—a 10 year follow-up study. Atherosclerosis 160:433–440
Ridker PM, Hennekens CH, Buring JE, Rifai N (2000) C-reactive protein and other markers of inflammation in the prediction of cardiovascular disease in women. N Engl J Med 342:836–843
Kushner I (1982) The phenomenon of the acute phase response. Ann N Y Acad Sci 389:39–48
Kushner I (2002) C-reactive protein and atherosclerosis. Science 297:520–521
Kimberly MM, Vesper HW, Caudill SP, Cooper GR, Rifai N, Dati F, Myers GL (2003) Standardization of immunoassays for measurement of high-sensitivity C-reactive protein. Phase I: evaluation of secondary reference materials. Clin Chem 49:611–616
Tanaka T, Horio T, Matuo Y (2002) Secretory production of recombinant human C-reactive protein in Escherichia Coli, capable of binding with phosphorylcholine, and its characterization. BBRC 295:163–166
Rzychon M, Zegers I, Schimmel H (2010) Analysis of the physicochemical state of C-reactive protein in different preparations including two certified reference materials. Clin Chem 56:1475–1482
Burkitt W, Pritchard C, Arsene C, Henrion A, Bunk D, O'Connor G (2008) Toward Systeme International d’Unite-traceable protein quantification: from amino acids to proteins. Anal Biochem 376:242–251
Kato M, Kato H, Eyama S, Takatsu A (2009) Application of amino acid analysis using hydrophilic interaction liquid chromatography coupled with isotope dilution mass spectrometry for peptide and protein quantification. J Chromatogr B 877:3059–3064
Kinumi T, Ichikawa R, Arimoto H, Takatsu A (2010) Traceable amino acid analyses of proteins and peptides by isotope-dilution mass spectrometry using precolumn derivatization reagent. Anal Sci 26:1007–1010
Jeong J, Lim H, Kim S, Ku H, Oh K, Park S (2011) Quantification of human growth hormone by amino acid composition analysis using isotope dilution liquid-chromatography tandem mass spectrometry. J Chromatogr A 1218:6596–6602
Munoz A, Kral R, Schimmel H (2011) Quantification of protein calibrants by amino acid analysis using isotope dilution mass spectrometry. Anal Biochem 408:124–131
Wu L, Yang B, Bi J, Wang J (2011) Development of bovine serum albumin certified reference. Anal Bioanal Chem 400:3443–3449
Pritchard C, Torma F, Hopley C, Quaglia M, O'Connor G (2011) Investigating microwave hydrolysis for the traceable quantification of peptide standards using gas chromatography-mass spectrometry. Anal Biochem 412:40–46
Kinumi T, Goto M, Eyama S, Kato M, Kasama T, Takatsu A (2012) Development of SI-traceable C-peptide certified reference material NMIJ CRM 6901-a using isotope-dilution mass spectrometry-based amino acid analyses. Anal Bioanal Chem 404:13–21
Eurachem guide (2012) Quantifying uncertainty in analytical measurements. LGC, Teddington
International Organization for Standardization Guide 35 (2006) Reference materials-general and statistical principles for certification, 3rd edn. ISO, Genova
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Published in the topical collection Reference Materials for Chemical Analysis with guest editors Hendrik Emons and Stephen A. Wise.
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Kato, M., Kinumi, T., Yoshioka, M. et al. Development of C-reactive protein certified reference material NMIJ CRM 6201-b: optimization of a hydrolysis process to improve the accuracy of amino acid analysis. Anal Bioanal Chem 407, 3137–3146 (2015). https://doi.org/10.1007/s00216-014-8190-0
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DOI: https://doi.org/10.1007/s00216-014-8190-0