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A novel automated indirect immunofluorescence autoantibody evaluation

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Abstract

Autoantibodies (AAb), especially antinuclear (ANAs) and anticytoplasmatic antibodies (ACyA), are essential diagnosing markers for several autoimmune diseases. The current gold standard method for ANA detection is manual indirect immunofluorescence (IIF) on human epithelial-2 (HEp-2) cells. However, this technique is cost and time consuming, and characterized by considerable intra- and interlaboratory variability. Thus, an automated IIF-HEp-2 reader has been developed recently. In the current study, we compared the performance of the automated AAb IIF-HEp-2 interpretation to conventional detection methods. Autoantibody detection by IIF on HEp-2 cells was performed in a total of 260 sera of patients, including 34 with systemic lupus erythematosus, 111 with dermatomyositis or polymyositis, 74 with systemic sclerosis, 41 with rare AAb patterns, and 137 healthy individuals. Visual interpretation and routine immunoassays were compared with a novel automated IIF-HEp-2 system using Aklides pattern recognition algorithms. Positive AAbs were detected in 95–100% of rheumatic patients by automated interpretation, in 74–100% with manual reading, and in 64–100% by immunodot assay. Receiver operating characteristic curve analysis of fluorescent intensity revealed a high sensitivity and specificity for automated reading of AAb with an agreement ranging from 90% to 95% between manual and automated interpretation (kappa 0.554–0.69) for systemic sclerosis and myositis, respectively. This study demonstrates a good correlation between manual and automated interpretation of AAb including ANA and ACyA in patients with autoimmune diseases. Full automation of HEp-2 cell assay reading may minimize errors in ANA pattern interpretation and thus help in the standardization of ANA assessment.

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Abbreviations

AAb:

Autoantibody

ANA:

Antinuclear antibody

ACyA:

Anticytoplasmatic antibody

IIF:

Indirect immunofluorescence

ELISA:

Enzyme-linked immunosorbent immunoassays

SLE:

Systemic lupus erythematosus

DM:

Dermatomyositis

PM:

Polymyositis

SSc:

Systemic sclerosis

Jo-1:

Histidyl-tRNA synthetase

PL-7:

Threonyl-tRNA synthetase

PL-12:

Alanyl-tRNA synthetase

SRP:

Signal recognition particle

ROC:

Receiver operating characteristics

MSA:

Mitotic spindle apparatus

AUC:

Area under curve

References

  1. Fritzler MJ (2008) Challenges to the use of autoantibodies as predictors of disease onset, diagnosis and outcomes. Autoimmun Rev 7:616–620

    Article  PubMed  CAS  Google Scholar 

  2. Sordet C, Goetz J, Sibilia J (2006) Contribution of autoantibodies to the diagnosis and nosology of inflammatory muscle disease. Joint Bone Spine 73:646–654

    Article  PubMed  CAS  Google Scholar 

  3. Worman HJ, Courvalin JC (2003) Antinuclear antibodies specific for primary biliary cirrhosis. Autoimmun Rev 2:211–217

    Article  PubMed  Google Scholar 

  4. Wiik A, Cervera R, Haass M, Kallenberg C, Khamashta M, Meroni PL, Piette JC, Schmitt R, Shoenfeld Y (2006) European attempts to set guidelines for improving diagnostics of autoimmune rheumatic disorders. Lupus 15:391–396

    Article  PubMed  CAS  Google Scholar 

  5. Lleo A, Invernizzi P, Gao B, Podda M, Gershwin ME (2010) Definition of human autoimmunity—autoantibodies versus autoimmune disease. Autoimmun Rev 9:A259–A266

    Article  PubMed  CAS  Google Scholar 

  6. Agmon-Levin N, Shapira Y, Selmi C, Barzilai O, Ram M, Szyper-Kravitz M, Sella S, Katz BS, Youinou P, Renaudineau Y, Larida B, Invernizzi P, Gershwin ME, Shoenfeld Y (2010) A comprehensive evaluation of serum autoantibodies in primary biliary cirrhosis. J Autoimmun 34:55–58

    Article  PubMed  CAS  Google Scholar 

  7. Arora-Singh RK, Assassi S, del Junco DJ, Arnett FC, Perry M, Irfan U, Sharif R, Mattar T, Mayes MD (2010) Autoimmune diseases and autoantibodies in the first degree relatives of patients with systemic sclerosis. J Autoimmun 35:52–57

    Article  PubMed  CAS  Google Scholar 

  8. Chretien P, Chousterman M, Abd Alsamad I, Ozenne V, Rosa I, Barrault C, Lons T, Hagege H (2009) Non-organ-specific autoantibodies in chronic hepatitis C patients: association with histological activity and fibrosis. J Autoimmun 32:201–205

    Article  PubMed  CAS  Google Scholar 

  9. Tozzoli R, Bizzaro N, Tonutti E, Villalta D, Bassetti D, Manoni F, Piazza A, Pradella M, Rizzotti P (2002) Guidelines for the laboratory use of autoantibody tests in the diagnosis and monitoring of autoimmune rheumatic diseases. Am J Clin Pathol 117:316–324

    Article  PubMed  Google Scholar 

  10. Hartung K, Seelig HP (2006) Laboratory diagnostics of systemic autoimmune diseases. Part 1. Collagenoses. Z Rheumatol 65:709–722, quiz 723–704

    Article  PubMed  CAS  Google Scholar 

  11. Wiik AS (2005) Anti-nuclear autoantibodies: clinical utility for diagnosis, prognosis, monitoring, and planning of treatment strategy in systemic immunoinflammatory diseases. Scand J Rheumatol 34:260–268

    Article  PubMed  CAS  Google Scholar 

  12. Nifli AP, Notas G, Mamoulaki M, Niniraki M, Ampartzaki V, Theodoropoulos PA, Kopnitsky MJ, Castanas E (2006) Comparison of a multiplex, bead-based fluorescent assay and immunofluorescence methods for the detection of ANA and ANCA autoantibodies in human serum. J Immunol Methods 311:189–197

    Article  PubMed  CAS  Google Scholar 

  13. Smith J, Onley D, Garey C, Crowther S, Cahir N, Johanson A, Painter S, Harradence G, Davis R, Swarbrick P (2005) Determination of ANA specificity using the UltraPlex platform. Ann N Y Acad Sci 1050:286–294

    Article  PubMed  CAS  Google Scholar 

  14. Ghirardello A, Bendo R, Rampudda ME, Bassi N, Zampieri S, Doria A (2009) Commercial blot assays in the diagnosis of systemic rheumatic diseases. Autoimmun Rev 8:645–649

    Article  PubMed  CAS  Google Scholar 

  15. Somers K, Govarts C, Stinissen P, Somers V (2009) Multiplexing approaches for autoantibody profiling in multiple sclerosis. Autoimmun Rev 8:573–579

    Article  PubMed  CAS  Google Scholar 

  16. Liu H, Norman GL, Shums Z, Worman HJ, Krawitt EL, Bizzaro N, Vergani D, Bogdanos DP, Dalekos GN, Milkiewicz P, Czaja AJ, Heathcote EJ, Hirschfield GM, Tan EM, Miyachi K, Bignotto M, Battezzati PM, Lleo A, Leung PS, Podda M, Gershwin ME, Invernizzi P (2010) PBC screen: an IgG/IgA dual isotype ELISA detecting multiple mitochondrial and nuclear autoantibodies specific for primary biliary cirrhosis. J Autoimmun 35:436–442

    Article  PubMed  CAS  Google Scholar 

  17. Lea P, Keystone E, Mudumba S, Kahama A, Ding SF, Hansen J, Azad AA, Wang S, Weber D (2009) Advantages of multiplex proteomics in clinical immunology: the case of rheumatoid arthritis: novel IgXPLEX: planar microarray diagnosis. Clin Rev Allergy Immunol 41(1):20–35

    Article  Google Scholar 

  18. Rigon A, Soda P, Zennaro D, Iannello G, Afeltra A (2007) Indirect immunofluorescence in autoimmune diseases: assessment of digital images for diagnostic purpose. Cytometry B Clin Cytom 72:472–477

    PubMed  Google Scholar 

  19. Egerer K, Roggenbuck D, Hiemann R, Weyer MG, Buttner T, Radau B, Krause R, Lehmann B, Feist E, Burmester GR (2010) Automated evaluation of autoantibodies on human epithelial-2 cells as an approach to standardize cell-based immunofluorescence tests. Arthritis Res Ther 12:R40

    Article  PubMed  Google Scholar 

  20. Hiemann R, Hilger N, Michel J, Nitschke J, Bohm A, Anderer U, Weigert M, Sack U (2007) Automatic analysis of immunofluorescence patterns of HEp-2 cells. Ann N Y Acad Sci 1109:358–371

    Article  PubMed  Google Scholar 

  21. Hiemann R, Buttner T, Krieger T, Roggenbuck D, Sack U, Conrad K (2009) Challenges of automated screening and differentiation of non-organ specific autoantibodies on HEp-2 cells. Autoimmun Rev 9:17–22

    Article  PubMed  CAS  Google Scholar 

  22. Wiik AS, Hoier-Madsen M, Forslid J, Charles P, Meyrowitsch J (2010) Antinuclear antibodies: a contemporary nomenclature using HEp-2 cells. J Autoimmun 35:276–290

    Article  PubMed  CAS  Google Scholar 

  23. Admou B, Essaadouni L, Amal S, Arji N, Chabaa L, El Aouad R (2009) Autoantibodies in systemic sclerosis: clinical interest and diagnosis approach. Ann Biol Clin (Paris) 67:273–281

    CAS  Google Scholar 

  24. Mierau R, Genth E (2006) Scleroderma associated autoantibodies—clinical and diagnostic relevance. Z Rheumatol 65:279–284

    Article  PubMed  CAS  Google Scholar 

  25. Tan EM (1989) Antinuclear antibodies: diagnostic markers for autoimmune diseases and probes for cell biology. Adv Immunol 44:93–151

    Article  PubMed  CAS  Google Scholar 

  26. Hachulla E, Launay D (2011) Diagnosis and classification of systemic sclerosis. Clin Rev Allergy Immunol 40(2):78–83

    Article  PubMed  Google Scholar 

  27. Solomon DH, Kavanaugh AJ, Schur PH (2002) Evidence-based guidelines for the use of immunologic tests: antinuclear antibody testing. Arthritis Rheum 47:434–444

    Article  PubMed  Google Scholar 

  28. Reshef T, Mekori YA, Mor A (2009) Differential antinuclear antibodies pattern. Isr Med Assoc J 11:123–124

    PubMed  Google Scholar 

  29. Meroni PL, Schur PH (2010) ANA screening: an old test with new recommendations. Ann Rheum Dis 69:1420–1422

    Article  PubMed  CAS  Google Scholar 

  30. Großmann K, Roggenbuck D, Schröder C, Conrad K, Schierack P, Sack U (2011) Multiplex assessment of non-organ specific autoantibodies with a novel microbead-based immunoassay. Cytometry A 79(2):118–125

    PubMed  Google Scholar 

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Disclosures

Dirk Roggenbuck has a management role and is a shareholder of GA Generic Assays GmbH and Medipan GmbH. Both companies are diagnostic manufacturers. All other authors declare that they have no competing financial interests.

Author information

Authors and Affiliations

  1. The Zabludowicz Center for Autoimmune Diseases, Tel-Aviv University, Tel-Aviv, Israel

    Shaye Kivity, Boris Gilburd, Nancy Agmon-Levin, Marina Garcia Carrasco, Yaron Tzafrir & Yehuda Shoenfeld

  2. Rheumatology Unit, Tel-Aviv University, Tel-Aviv, Israel

    Shaye Kivity

  3. Department of Medicine ‘A & C’, Tel-Aviv University, Tel-Aviv, Israel

    Shaye Kivity

  4. Department of Medicine ‘A & C’, Sheba Medical Center, Tel Hashomer, Israel

    Yael Sofer

  5. Blood Center, Tel-Aviv University, Tel-Aviv, Israel

    Matilda Mandel

  6. Department of Medicine B, Sheba Medical Center, Tel-Hashomer, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv, Israel

    Yehuda Shoenfeld

  7. Department of Rheumatology, Clinical Hospital Buenos Aires University, Buenos Aires, Argentina

    Marina Garcia Carrasco

  8. GA Generic Assays GmbH, Dahlewitz, Berlin, Germany

    Thomas Buttner & Dirk Roggenbuck

  9. Department of Biomedicine, Division of Rheumatology AOUC, Villa Monna Tessa, Viale Pieraccini 18, Florence, 50139, Italy

    Marco Matucci-Cerinic

  10. Medical and Health Science Center, University of Debrecen, Debrecen, 4012, Hungary

    Katalin Danko

  11. Servicio Inmunología Hospital Universitario Marqués de aldecilla, 39008, Santander, Spain

    Marcos López Hoyos

  12. Incumbent of the Laura Schwarz-Kip Chair for Research of Autoimmune Diseases, Tel-Aviv University, Tel-Aviv, Israel

    Yehuda Shoenfeld

Authors
  1. Shaye Kivity
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  2. Boris Gilburd
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  3. Nancy Agmon-Levin
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  4. Marina Garcia Carrasco
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  5. Yaron Tzafrir
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  6. Yael Sofer
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  7. Matilda Mandel
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  8. Thomas Buttner
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  9. Dirk Roggenbuck
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  10. Marco Matucci-Cerinic
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  11. Katalin Danko
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  12. Marcos López Hoyos
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  13. Yehuda Shoenfeld
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Corresponding author

Correspondence to Yehuda Shoenfeld.

Additional information

Shaye Kivity and Boris Gilburd have equally contributed.

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Kivity, S., Gilburd, B., Agmon-Levin, N. et al. A novel automated indirect immunofluorescence autoantibody evaluation. Clin Rheumatol 31, 503–509 (2012). https://doi.org/10.1007/s10067-011-1884-1

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  • DOI: https://doi.org/10.1007/s10067-011-1884-1

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