Quantification of Lupus Anticoagulants in Clinical Samples Using Anti- β2GP1 and Anti-Prothrombin Monoclonal Antibodies

Summary

Quantification of lupus anticoagulant (LA) in clinical samples is hampered by the lack of a suitable standard of activity. We evaluated the use of mAbs displaying LA activity for this purpose. As most patient samples contain both β₂Glycoprotein I (β₂GP1) and prothrombin dependent LA, a combination of two mAbs, one of each specificity, was added to normal plasma in a concentration from 0 to 60 g/ml. Eight assay systems using different reagents and instruments were used. The calibration curves were linear for all but one, with marked differences between the responsiveness to each mAb. A panel of plasmas from 69 patients with persistent LA diagnosed using the SSCISTH criteria was tested. An antiphospholipid syndrome (APS) was present in 40, whereas 29 were asymptomatic. LA activities of individual plasmas varied between assays (p <10^–4), but homogeneous subgroups were identified. In a majority of samples, LA activity displayed a prothrombin-dependent profile, with a variable contribution of β₂GP1-dependent activity. The latter was associated to β₂GP1 antibodies detected by solid-phase immunoassay. By using 3 dilute Russell viper venom time assays, higher LA titers were found in APS, compared to asymptomatic patients (p <0.05).

Keywords

Lupus anticoagulant - anti-β₂glycoprotein 1 - anti-prothrombin - monoclonal antibodies - antiphospholipid syndrome

• References

• 1 Bevers EM. Lupus Anticoagulant IgG’s (LA) are not directed to phospholipids only, but to a complex of lipid-bound human prothrombin. Thromb Haemost 1991; 66: 629-32.
• 2 Oosting JD, Derksen R, Entjes HT, Bouma BN, De Groot PG. Lupus anticoagulant activity is frequently dependent on the presence of β₂ Glyco-protein I. Thromb Haemost 1992; 67: 499-502.
• 3 Galli M, Comfurius P, Barbui T, Zwaal RFA, Bevers EM. Anticoagulant Activity of β₂-Glycoprotein I is Potentiated by a Distinct Subgroup of Anti-cardiolipin Antibodies. Thromb Haemost 1992; 68: 297-300.
• 4 Horbach DA, Van Oort E, Derksen RHWM, De Groot PG. The Contribution of Anti-prothrombin-antibodies to Lupus Anticoagulant Activity. Thromb Haemost 1998; 79: 790-5.
• 5 Arvieux J, Regnault V, Hachulla E, Darnige L, Roussel B, Bensa JC. Heterogeneity and Immunochemical Properties of Anti-β₂-glycoprotein I autoantibodies. Thromb Haemost 1998; 80: 393-8.
• 6 Akimoto T, Akama T, Kono I, Sumida T. Relationship between clinical features and binding domains of anti-prothrombin autoantibodies in patients with systemic lupus erythematosus and antiphospholipid syndrome. Lupus 1999; 8: 761-6.
• 7 Ginsberg JS, Wells PS, Brill-Edwards P, Donovan D, Moffatt K, Johnston M. et al. Antiphospholipid Antibodies and Venous Thromboembolism. Blood 1995; 86: 3685-91.
• 8 Horbach DA. Lupus Anticoagulant is the Strongest Risk Factor for both Venous and Arterial Thrombosis in Patients with Systemic Lupus Erythematosus. Thromb Haemost 1996; 76: 916-24.
• 9 Wahl DG. et al. Risk for venous thrombosis related to antophospholipid antibodies in systemic lupus erythematosus: a meta-analysis. Lupus 1997; 6: 467-73.
• 10 Wahl DG, Guillemin F, de Maistre E, Perret-Guillaume C, Lecompte T, Thibaut G. Meta-analysis of the risk of venous thrombosis in individuals with antiphospholipid antibodies without underlying autoimmune disease or previous thrombosis. Lupus 1998; 7: 15-22.
• 11 Galli M, Finazzi G, Bevers EM, Barbui T. Kaolin Clotting Time and Dilute Russell’s Viper Venom Time Distinguish Between Prothrombin dependent and β₂-Glycoprotein I-Dependent Antiphospholipid Antibodies. Blood 1995; 86: 617-23.
• 12 Pengo V, Biasiolo A, Brocco T, Tonett S, Ruffatti A. Autoantibodies to Phospholipid-binding Plasma Proteins in Patients with Thrombosis and Phospholipid-reactive Antibodies. Thromb Haemost 1996; 75: 721-4.
• 13 Swadzba J, Declerck LS, Stevens WJ. Association of Anticardiolipin anti- β₂ Glycoprotein antiprothrombin antibodies, and lupus anticoagulants in patients with systemic lupus erythematosus with a history of thrombosis. J Rhumatol 1997; 24: 1710-5.
• 14 D’Angelo A, Safa O, Crippa C, Garlando A, Sabbadini MG, Vigano D’Angelo S. Relationship of lupus anticoagulant, anticardiolipin, anti-β_2-GPI and anti-prothrombin autoantibodies with history of thrombosis in patients with the clinical suspicion of APA-syndrome. Thromb Haemost 1997; 78: 697-968.
• 15 Galli M, Barbui T. Anti-prothrombin Antibodies: Detection and Clinical significance in the Antiphospholipid syndrome. Blood 1999; 93: 2149-57.
• 16 Arvieux J, Pouzol P, Roussel B, Jacob MC, Colomb MG. Lupus-like anticoagulant properties of murine antibodies to β₂-glycoprotein I. Br J Haematol 1992; 81: 568-73.
• 17 Arnout J, Wittevrongel C, Vanrusselt M, Hoylaerts M, Vermylen J. β₂-glycoprotein I dependent lupus anticoagulants from stable bivalent antibody β₂-glycoprotein I complexes on phospholipid surface. Thromb Haemost 1998; 79: 79-86.
• 18 Arnout J, Vanrussel M, Wittevrongel C, Vermylen J. Monoclonal Antibodies against β₂-Glycoprotein I: Use as Reference Material for Lupus Anticoagulant Tests. Thromb Haemost 1998; 79: 955-8.
• 19 Arnout J, Meijer P, Vermylen J. Lupus Anticoagulant Testing in Europe: An analysis of Results from the First European Concerted Action on Thrombophilia (ECAT) Survey Using Plasmas Spiked with Monoclonal Antibodies against Human β₂-Glycoprotein I. Thromb Haemost 1999; 81: 929-34.
• 20 Arnout J, Vanrusselt M, Nevens C, Smans K, Wittevrongel C, Vermylen J. Some Murine Monoclonal Antibodies Against Human Prothrombin have Lupus Anticoagulant Activity. Thromb Haemost. 1999 Suppl August: 65 (abstract).
• 21 Lawrie AS, Mackie IJ, Purdy G, Machin SJ. Effect of Instruments on Lupus Anticoagulant Testing – Rebuttal. Thromb Haemost 1999; 83: 346-8.
• 22 Goudemand J, Caron C, De Prost D, Derlon A, Borg JY, Sampol J. et al. Evaluation of Sensitivity and Specificity of a Standardized Procedure Using different Reagents for the Detection of Lupus Anticoagulants. Thromb Haemost 1997; 77: 336-42.
• 23 Lawrie AS, Mackie IJ, Purdy G, Machin SJ. The Sensitivity and Specificity of Commercial Reagents for the detection of Lupus Anticoagulant Show Marked Differences in Performance between Photo-optical and Mechanical Coagulometers. Thromb Haemost 1999; 81: 758-62.
• 24 Triplett DA, Brandt JT, Maas RL. The laboratory heterogeneity of Lupus Anticoagulants. Arch Pathol Lab Med 1985; 109: 946-51.
• 25 Brandt JT, Triplett DA, Alving B, Scharrer I. Criteria for the diagnosis of Lupus Anticoagulants: an update. Thromb Haemost 1995; 74: 1185-90.
• 26 Harris EN. Antiphospholipids antibodies. Br J Haematol 1990; 74: 1-9.
• 27 Jacobsen EM, Barna-Cler L, Taylor JM, Triplett DA, Wisloff F. The Lupus Ratio Test – An Interlaboratory Study on the detection of Lupus Anticoagulants by an APTT-based Integrated, and Semi-quantitative Test. Thromb Haemost 2000; 83: 704-8.
• 28 Brandt JT, Barna K, Triplett DA. Laboratory identification of lupus anticoagulants: results of the second international workshop for identification of lupus anticoagulants. Thromb Haemost 1995; 74: 1597-603.
• 29 Arnout J, Vanrusselt M, Nevens C, Wittevrongel C, Vermylen J. Interaction between β₂GP1- and Prothrombin-Dependent Lupus Anticoagulants. Thromb Haemost. 1999 supp August: 70 (abstract).
• 30 Pengo V, Biasiolo A, Rampazzo P, Brocco T. dRVVT is more sensitive than KCT or TTI for Detecting Lupus Anticoagulant Activity of Anti-β_2-glycoprotein I Autoantibodies. Thromb Haemost 1999; 81: 256-8.
• 31 Norbis F, Barbui T, Galli M. Comparison of two Russell’s viper venom time (dRVVT) for distinguishing the phospholipid-dependent inhibitors of coagulation. Thromb Haemost. 1997 Suppl June: 331 (Abstract).
• 32 Callahan JB, Manares CC, Moll S, Ortel TL. Inability to distinguish between thrombotic and non-thrombotic antiphospholipid antibody positive patients with kaolin clotting time and dilute Russell‘s viper venom assay ratio. Lupus 1996; 5: 53 (abstract).
• 33 Rivard GE, Schiffman S, Rapaport SI. Cofactor of the lupus anticoagulant. Thrombosis and Diathesis Haemorragica 1974; 32: 554-63.
• 34 Galli M, Finazzi G, Norbis F, Marziali S, Marchioli R, Barbui T. The Risk of Thrombosis in Patients with Lupus Anticoagulants is Predicted by their Specific Coagulation Profile. Thromb Haemost 1999; 81: 695-700.
• 35 Brandt JT, Triplett DA. The effect of phospholipids on the detection of Lupus Anticoagulants by the dilute Russell Viper Venom Time. Arch Pathol Lab Med 1989; 113: 1376-8.
• 36 Arvieux J, Darnige L, Caron C, Reber G, Bensa JC, Colomb MG. Development of an ELISA for autoantibodies to prothrombin showing their prevalence in patients with lupus anticoagulant. Thromb Haemost 1995; 74: 1120-5.
• 37 Pengo V, Brocco T, Biasiolo A, Rampazzo P, Carraro P, Zamarchi R. Procoagulant Effect of Anti-β₂-Glycoprotein I Antibodies With Lupus Anticoagulant Activity. Blood 1999; 94: 3814-9.
• 38 Galli M, Ruggeri L, Barbui T. Differential Effects of Anti-β₂-Glycoprotein I and Antiprothrombin Antibodies on the Anticoagulant Activity of Activated Protein C. Blood 1998; 91: 1999-2004.
• 39 Exner T, Hohnen-Behrens C. Modified Clotting Tests For Subtyping “Lupus” Anticoagulants And The Use Of Artificial Standards. Thromb Haemost. 1999 supp August: 768 (abstract).
• 40 Galli M, Diott J, Norbis F, Ruggeri L, Cler L, Triplett DA, Barbui T. Lupus anticoagulants and Thrombosis: Clinical Association of Different Coagulation and Immunologic Tests. Thromb Haemost 2000; 84: 1012-6.