Distinguishing Plasma Lupus Anticoagulants from Anti-Factor Antibodies Using Hexagonal (II) Phase Phospholipids

Abstract

SummaryThe association of lupus anticoagulant antibodies with thrombosis, thrombocytopenia, and multiple spontaneous abortions underlines the importance of diagnostic assays which are able to distinguish these antibodies from anti-factor antibodies and factor deficiencies, as all three prolong in vitro coagulation assays measuring activated partial thromboplastin time (APTT). Heparin also prolongs the APTT assay and often interferes with the detection of lupus anticoagulant activity. The present study describes a direct and simple dilute APTT assay in which plasma is preincubated with hexagonal (II) phase phosphatidyl ethanolamine (PE). Using this system, the lupus anticoagulant antibody activity of 10 randomly selected plasmas from SLE patients was inhibited by 81.2-99.5%, while prolongation of the APTT assay by 6 different anti-factor antibody-containing plasmas, 5 factor deficient plasmas, and 6 heparin-containing plasmas remained unaffected. Inhibition was dependent on epitopes exposed when PE was presented in the hexagonal (II) phase. This data suggests that hexagonal (II) PE is specifically recognized by lupus anticoagulant antibodies in SLE patients and may play a role in the etiology of the disease.

Echocardiographic Abnormalities and Antiphospholipid Antibodies in Patients with Systemic Lupus Erythematosus

“…In the tests considered to show alteration, the presence of the lupus anticoagulant as a coagulation inhibitor and a cause of the increase in coagulation time was confirmed through the reagent with a higher concentration of phospholipids [16][17][18] . A ratio between the time using the low-phospholipid reagent and that using the high-phospholipid reagent equal to or greater than 1.2 was considered positive.…”

The epitopes for some antiphospholipid antibodies are adducts of oxidized phospholipid and β2glycoprotein 1 (and other proteins)

“…Rauch, Janoff, and colleagues proposed that ␤ 2 GP1-induced changes in the three-dimensional structure of cardiolipin (CL) were key to the PL functioning as an antigen in conventional aCL immunoassays and to the immunogenicity of the PL in vivo (11)(12)(13). Others also have shown that binding of some aCLs is dependent on interactions between PL and PL-binding proteins (14)(15)(16)(17).…”

Immunization with an Apoptotic Cell-Binding Protein Recapitulates the Nephritis and Sequential Autoantibody Emergence of Systemic Lupus Erythematosus

“…Platelet poor plasma was isolated by centrifugation for 3 min at 11,600 ϫ g in a tabletop microfuge (MicroCentaur; Accurate Chemical and Scientific). Murine plasma was diluted 1/1 with citrated normal human plasma and was tested for lupus anticoagulant activity in a dilute activated partial thromboplastin time (aPTT) assay, as previously described (19,20). aPTT values were considered significantly prolonged if they exceeded the mean plus 2 SE of the aPTT value of plasma from control mice immunized with HEPES or cells plus HEPES.…”