Background: Fully automated chemiluminescence immunoassays (CLIAs) are emerging technologies for the detection of anti-cardiolipin (aCL) and anti-β2 glycoprotein I (anti-β2GPI) antibodies for anti-phospholipid syndrome (APS) classification, which is commonly based on an enzyme-linked immunosorbent assay (ELISA) test result. CLIA and a homemade ELISA were used in this study to detect these antibodies, and their performances were compared.
Methods: Sera were collected from 104 patients with primary APS, 88 seronegative subjects who met the clinical but not the laboratory criteria for APS, and 150 control subjects. IgG/IgM aCL and IgG/IgM anti-β2GPI antibodies were determined in the sera using a CLIA (HemosIL AcuStar®) and a homemade ELISA.
Results: CLIA had a significantly lower comparative sensitivity for IgM aCL and IgG/IgM IgG anti-β2GPI antibodies; its comparative specificity was higher with respect to ELISA for IgM aCL and IgM anti-β2GPI antibodies. The two techniques showed a high, significant agreement (p<0.001) and a significant titer correlation (p<0.001). CLIA also detected IgG/IgM aCL and IgG anti-β2GPI antibodies in the seronegative patients. There was a significantly higher prevalence of IgG aCL and IgG anti-β2GPI antibodies (p<0.001 and p=0.01, respectively) in those patients with respect to that in the control population.
Conclusions: Despite a lower comparative sensitivity, CLIA showed a higher comparative specificity for some aPL and a good level of agreement and correlation with a homemade ELISA. CLIA also detected some aCL and anti-β2GPI antibodies in the seronegative patients not usually identified by homemade ELISA.
The authors are grateful to Dr Elisa Salvan for the statistical calculations and to Mrs Linda Inverso Moretti for editing the English version of the manuscript.
Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.
Financial support: None declared.
Employment or leadership: None declared.
Honorarium: None declared.
Competing interests: The funding organization(s) played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.
1. Miyakis S, Lockshin MD, Atsumi T, Branch DW, Brey RL, Cervera R, et al. International consensus statement on an update of the classification criteria for definite antiphospholipid syndrome (APS). J Thromb Haemost 2006;4:295–306.10.1111/j.1538-7836.2006.01753.xSearch in Google Scholar
4. Pengo V, Tripodi A, Reber G, Rand JH, Ortel TL, Galli M, et al. Update of the guidelines for lupus anticoagulant detection. Subcommittee on Lupus Anticoagulant/Antiphospholipid Antibody of the Scientific and Standardisation Committee of the International Society on Thrombosis and Haemostasis. J Thromb Haemost 2009;7:1737–40.10.1111/j.1538-7836.2009.03555.xSearch in Google Scholar
5. Keeling D, Mackie I, Moore GW, Greer IA, Greaves M. British Committee for Standards in Haematology Guidelines on the investigation and management of antiphospholipid syndrome. Br J Haematol 2012;157:47–58.10.1111/j.1365-2141.2012.09037.xSearch in Google Scholar
6. Clinical and Laboratory Standards Institute (CLSI). Laboratory testing for the lupus anticoagulant; approved Guideline. CLSI document H60-A. Wayne, PA: CLSI, 2014.Search in Google Scholar
7. Favaloro EJ, Wong RC. Antiphospholipid antibody testing for the antiphospholipid syndrome: a synopsis of challenges and recent guidelines. Pathology 2014;46:481–95.Search in Google Scholar
9. Reber G, Tincani A, Sanmarco M, de Moerloose P, Boffa MC. Proposals for the measurement of anti-b2-glycoprotein I antibodies. Standardization Group of the European Forum on Antiphospholipid Antibodies. J Thromb Haemost 2004;2:1860–2.10.1111/j.1538-7836.2004.00910.xSearch in Google Scholar
10. Tincani A, Allegri F, Balestrieri G, Reber G, Sanmarco M, Meroni P, et al. Minimal requirements for antiphospholipid antibodies ELISAs proposed by the European Forum on antiphospholipid antibodies. Thromb Res 2004;114:553–8.10.1016/j.thromres.2004.06.035Search in Google Scholar
11. Wong RC, Gillis D, Adelstein S, Baumgart K, Favaloro EJ, Hendle MJ, et al. Consensus guidelines on anti-cardiolipin antibody testing and reporting. Pathology 2004;36:63–8.10.1080/00313020310001643615Search in Google Scholar
12. Wong RC, Favaloro EJ, Adelstein S, Baumgart K, Bird R, Brighton TA, et al. Consensus guidelines on anti-beta 2 glycoprotein I testing and reporting. Pathology 2008;40:58–63.10.1080/00313020701717720Search in Google Scholar
14. Lakos G, Favaloro EJ, Harris EN, Meroni PL, Tincani A, Wong RC, et al. International consensus guidelines on anticardiolipin and anti-β2-glycoprotein I testing: report from the 13th International Congress on Antiphospholipid Antibodies. Arthritis Rheum 2012;64:1–10.10.1002/art.33349Search in Google Scholar
15. Pierangeli SS, Favaloro EJ, Lakos G, Meroni PL, Tincani A, Wong RC. Standards and reference materials for the anticardiolipin and anti-β2glycoprotein I assays: a report of recommendations from the APL Task Force at the 13th International Congress on Antiphospholipid Antibodies. Clin Chim Act 2012;413:358–60.10.1016/j.cca.2011.09.048Search in Google Scholar
16. Pierangeli SS, Harris EN. A quarter of a century in anticardiolipin antibody testing and attempted standardization has led us to here, which is? Semin Thromb Hemost 2008;34:313–28.10.1055/s-0028-1085473Search in Google Scholar
17. Reber G, Boehlen F, de Moerloose P. Technical aspects in laboratory testing for antiphospholipid antibodies: is standardization an impossible dream? Semin Thromb Hemost 2008;34:340–6.10.1055/s-0028-1085476Search in Google Scholar
18. Wong R, Favaloro E, Pollock W, Wilson R, Hendle M, Adelstein S. A multi-centre evaluation of the intra-assay and inter-assay variation of commercial and in-house anti-cardiolipin antibody assays. Pathology 2004;36:182–92.10.1080/00313020410001672037Search in Google Scholar
21. Baeyens WR, Schulman SG, Calokerinos AC, Zhao Y, García Campaña AM, Nakashima K, et al. Chemiluminescence-based detection: principles and analytical applications in flowing streams and in immunoassays. J Pharm Biomed Anal 1998;17:941–53.10.1016/S0731-7085(98)00062-4Search in Google Scholar
22. Andreoli L, Rizzini S, Allegri F, Meroni P, Tincani A. Are the current attempts at standardization of antiphospholipid antibodies still useful? Emerging technologies signal a shift in direction. Semin Thromb Hemost 2008;34:356–60.10.1055/s-0028-1085478Search in Google Scholar
23. De Moerloose P, Reber G, Musial J, Arnout J. Analytical and clinical performance of a new, automated assay panel for the diagnosis of antiphospholipid syndrome. J Thromb Haemost 2010;8:1540–6.10.1111/j.1538-7836.2010.03857.xSearch in Google Scholar
24. Persijn L, Decavele AS, Schouwers S, Devreese K. Evaluation of a new set of automated chemiluminescense assays for anticardiolipin and anti-beta2-glycoprotein I antibodies in the laboratory diagnosis of the antiphospholipid syndrome. Thromb Res 2011;128:565–9.10.1016/j.thromres.2011.04.004Search in Google Scholar
25. Van Hoecke F, Persijn L, Decavele AS, Devreese K. Performance of two new, automated chemiluminescence assay panels for anticardiolipin and anti-beta2-glycoprotein I antibodies in the laboratory diagnosis of the antiphospholipid syndrome. Int J Lab Hematol 2012;34:630–40.10.1111/j.1751-553X.2012.01448.xSearch in Google Scholar
26. Capozzi A, Lococo E, Grasso M, Longo A, Garofalo T, Misasi R, et al. Detection of antiphospholipid antibodies by automated chemiluminescence assay. J Immunol Methods 2012;379:48–52.10.1016/j.jim.2012.02.020Search in Google Scholar
27. Forastiero R, Papalardo E, Watkins M, Nguyen H, Quirbach C, Jaskal K, et al. Evaluation of different immunoassays for the detection of antiphospholipid antibodies: report of a wet workshop during the 13th International Congress on Antiphospholipid Antibodies. Clin Chim Acta 2014;428:99–105.10.1016/j.cca.2013.11.009Search in Google Scholar
28. Mondejar R, González-Rodríguez C, Toyos-Sáenz de Miera FJ, Melguizo-Madrid E, Zohoury N, Mahler M. Role of antiphospholipid score and anti-β2-glycoprotein I Domain I autoantibodies in the diagnosis of antiphospholipid syndrome. Clin Chim Acta 2014;431:174–8.10.1016/j.cca.2014.01.047Search in Google Scholar
29. Chung Y, Kim JE, Lim HS, Kim HK. Clinical performance of anticardiolipin and antiβ2 glycoprotein I antibodies using a new automated chemiluminescent assay: superior thrombotic prediction of combined results measured by two different methods. Blood Coagul Fibrinolysis 2014;25:10–5.10.1097/MBC.0b013e32836466b5Search in Google Scholar
30. Ichikawa K, Tsutsumi A, Atsumi T, Matsuura E, Kobayashi S, Hughes GR, et al. A chimeric antibody with the human gamma1 constant region as a putative standard for assays to detect IgG beta2-glycoprotein I-dependent anticardiolipin and anti-beta2-glycoprotein I antibodies. Arthritis Rheum 1999;42:2461–70.10.1002/1529-0131(199911)42:11<2461::AID-ANR25>3.0.CO;2-OSearch in Google Scholar
31. Ruffatti A, Olivieri S, Tonello M, Bortolati M, Bison E, Salvan E, et al. Influence of different IgG anticardiolipin antibody cut-off values on antiphospholipid syndrome classification. J Thromb Haemost 2008;6:1693–6.10.1111/j.1538-7836.2008.03121.xSearch in Google Scholar
32. Viera AJ, Garrett JM. Understanding interobserver agreement: the kappa statistic. Fam Med 2005;37:360–3.Search in Google Scholar
33. Fontana P, Poncet A, Lindhoff-Last E, de Moerloose P, Devreese KM. Refinement of the cutoff values of the HemosIL AcuStar assay for the detection of anticardiolipin and anti-beta2-glycoprotein-1 antibodies. J Thromb Haemost 2014;12:2034–7.10.1111/jth.12732Search in Google Scholar
34. Galli M, Luciani D, Bertolini G, Barbui T. Lupus anticoagulants are stronger risk factors for thrombosis than anticardiolipin antibodies in the antiphospholipid syndrome: a systematic review of the literature. Blood 2003;101:1827–32.10.1182/blood-2002-02-0441Search in Google Scholar
35. Galli M, Reber G, de Moerloose P, de Groot PG. Invitation to a debate on the serological criteria that define the antiphospholipid syndrome. J Thromb Haemost 2008;6:399–401.10.1111/j.1538-7836.2008.02862.xSearch in Google Scholar
36. Galli M, Luciani D, Bertolini G, Barbui T. Anti-beta 2-glycoprotein I, antiprothrombin antibodies, and the risk of thrombosis in the antiphospholipid syndrome. Blood 2003;102:2717–23.10.1182/blood-2002-11-3334Search in Google Scholar
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