Species Specificity of Anti-β₂ Glycoprotein I Autoantibodies and Its Relevance to Anticardiolipin Antibody Quantitation

Summary

Some patients suspected of having antiphospholipid antibody syndrome (APS) were found to be positive for anti-β₂ glycoprotein I (β₂GPI) antibodies despite negative results for antibodies to cardiolipin (ACA). Since the major source of β₂GPI in the ACA assay is animal (usually bovine) serum, we studied the influence on ACA quantitation of the species specificity of anti-β₂GPI antibodies from patients with various autoimmune disorders, mostly systemic lupus erythematosus and primary APS. Ninety-seven sera were selected based on IgG (n = 76) or IgM (n = 64) positivity by ELISA using γ-irradiated plates coated with human or bovine purified β₂GPI. A higher proportion of IgM (43.7%) than IgG (7.9%) reacted to human, but not bovine, β₂GPI. Furthermore, from the samples reactive to both proteins, the ratio of antibody level against bovine to that against human β₂GPI was 1.08 ± 0.58 for IgG and 0.58 ± 0.3 for IgM (p <10⁻⁵). IgG and IgM ACA were detected in 78 and 40 sera, respectively; concordance between the two ELISAs for ACA and anti-β₂GPI antibodies was 94% for IgG and 75% for IgM. Out of 28 IgM showing recognition restricted to human β₂GPI, 21 were missed by the ACA assay, possibly because of lower concentrations of β₂GPI in those patients’ sera. The antibody reactivity pattern towards human and bovine β₂GPI of individual sera showed no variation with time and was related to the relative antibody avidity for each protein. A murine anti-human β₂GPI monoclonal antibody, 9G1, that cross-reacts with bovine β₂GPI, competed to a large extent with the patients’ anti-β₂GPI antibody binding sites whatever isotype involved or protein recognized. Therefore, anti-β₂GPI antibodies of IgM isotype display a marked preference for human compared to bovine β₂GPI responsible for frequent inconsistencies in the ACA assay.

• References

• 1 Triplett DA. Antiphospholipid-protein antibodies: laboratory detection and clinical relevance. Thromb Res 1995; 78: 1-31
• 2 Harris EN, Gharavi AE, Patel SP, Hughes GRV. Evaluation of the anti-cardiolipin antibody test: report of an international workshop held 4 April 1986. Clin Exp Immunol 1987; 68: 215-222
• 3 Harris EN, Pierangeli S, Birch D. Anticardiolipin wet workshop report: fifth international symposium on antiphospholipid antibodies. Am J Clin Pathol 1994; 101: 616-624
• 4 Matsuura E, Igarashi M, Igarashi Y, Nagae H, Ichikawa K, Yasuda T, Koike T. Molecular definition of human ²₂-glycoprotein I (²₂-GPI) by cDNA cloning and inter-species differences of ²₂-GPI in alternation of anticardiolipin binding. Int Immunol 1991; 3: 1217-1221
• 5 Matsuura E, Igarashi Y, Nagae H, Yasuda T, Triplett DA, Koike T. Anticardiolipin antibodies recognize ²₂-glycoprotein I structure altered by interacting with an oxygen modified solid phase surface. J Exp Med 1994; 179: 457-462
• 6 Roubey RAS. Autoantibodies to phospholipid-binding plasma proteins: a new view of lupus anticoagulants and other “antiphospholipid” autoantibodies. Blood 1994; 84: 2854-2867
• 7 Arvieux J, Roussel B, Jacob MC, Colomb MG. Measurement of antiphospholipid antibodies by ELISA using ²₂-glycoprotein I as an antigen. J Immunol Meth 1991; 143: 223-229
• 8 Pullen GR, Fitzgerald MG, Hosking CS. Antibody avidity determination by ELISA using thiocyanate elution. J Immunol Meth 1986; 86: 83-87
• 9 Arvieux J, Pouzol P, Roussel B, Jacob MC, Colomb MG. Lupus-like anticoagulant properties of murine monoclonal antibodies to ²₂-glycoprotein I. BrJ Haematol 1992; 81: 568-573
• 10 Reber G, Arvieux J, Comby E, Degenne D, de Moerloose P, Sanmarco M, Potron G. Multicenter evaluation of nine commercial kits for the quantitation of anticardiolipin antibodies. Thromb Haemost 1995; 73: 444-452
• 11 Sammaritano LR, Lockshin MD, Gharavi AE. Antiphospholipid antibodies differ in aPL cofactor requirement. Lupus 1992; 1: 83-90
• 12 McCarthy JM, Wagenknecht DR, McIntyre JA. Activity of antiphospholipid antibody ELISA cofactor in different animal sera. J Clin Lab Anal 1994; 8: 167-171
• 13 Kertesz Z, Yu B, Steinkasserer A, Haupt H, Benham A, Sim RB. Characterization of binding of human ²₂-glycoprotein I to cardiolipin. Biochem J 1995; 310: 315-321
• 14 Kamboh MI, Wagenknecht DR, McIntyre JA. Heterogeneity of the apo-lipoprotein H*3 allele and its role in affecting the binding of apolipopro-tein H (²₂-glycoprotein I) to anionic phospholipids. Hum Genet 1995; 95: 385-388
• 15 Benjamin DC, Berzofsky JA, East IJ, Gurd FRN, Hannum C, Leach SJ, Margoliash E, Michael JG, Miller A, Prager EM, Reichlin M, Sercarz EE, Smith-Gill SJ, Todd PE, Wilson AC. The antigenic structure of proteins: a reappraisal. Ann Rev Immunol 1984; 2: 67-101
• 16 Hunt JE, Krilis SA. The fifth domain of ²₂-glycoprotein I contains a phospholipid binding site (Cys281-Cys288), and a region recognized by anticardiolipin antibodies. J Immunol 1994; 152: 653-659
• 17 Wang M, Kandiah DA, Ichikawa K, Khamashta M, Hughes G, Koike T, Roubey R, Krilis SA. Epitope specificity of monoclonal anti-²₂-glycoprotein I antibodies derived from patients with the antiphospholipid syndrome. J Immunol 1995; 155: 1629-1636
• 18 Nomura M, Okada J, Kondo H. Detection of anti-bovine ²₂-glycoprotein I antibody in sera from patients with antiphospholipid syndrome. Rheumatol Int 1995; 15: 1-8
• 19 Arvieux J, Roussel B, Ponard D, Colomb MG. IgG2 subclass restriction of anti-²₂- glycoprotein I antibodies in autoimmune patients. Clin Exp Immunol 1994; 95: 310-315
• 20 Kouts S, Wang M, Adelstein S, Krilis SA. Immunization of a rabbit with ²₂-glycoprotein I induces charge-dependent crossreactive antibodies that bind anionic phospholipids and have similar reactivity as autoimmune antiphospholipid antibodies. J Immunol 1995 155: 958-966
• 21 Cabiedes J, Cabral A, Alarcon-Segovia D. Clinical manifestations of the antiphospholipid syndrome in patients with systemic lupus erythematosus associate more strongly with anti-²₂-glycoprotein I than with antiphospholipid antibodies. J Rheumatol 1995; 22: 1899-1906
• 22 Arvieux J, Damige L, Caron C, Reber G, Bensa JC, Colomb MG. Development of an ELISA for autoantibodies to prothrombin showing their prevalence in patients with lupus anticoagulants. Thromb Haemost 1995; 74: 1120-1125