Development of an ELISA for autoantibodies to prothrombin showing their prevalence in patients with lupus anticoagulants.
Some lupus anticoagulants (LA) have been shown to be directed against phospholipid-bound prothrombin. While developing an ELISA to detect anti-prothrombin autoantibodies in patient serum or plasma, no or very low signal was observed using human prothrombin immobilized on plain polystyrene plates. In contrast, the same LA-positive samples bound specifically to prothrombin coated on gamma-irradiated plates, depending on the radiation dose, in the absence of added calcium and phospholipid. Optimization of the assay required the addition of 0.1% Tween 20 to the buffers. Antibody specificity for immobilized prothrombin was ascertained by competition using liposome-bound prothrombin, since fluid-phase prothrombin competed poorly. Seventy-seven of 139 patients (55.4%) with LA related to a variety of underlying diseases possessed anti-prothrombin antibodies (27 IgG, 35 IgM and 15 both isotypes), either isolated or more often associated with anti-beta 2 glycoprotein I (beta 2GPI) antibodies. These included 67-71% of the patients with systemic lupus erythematosus and related disorders, primary antiphospholipid antibody syndrome or drug-induced LA (autoimmune groups), but only 19-20% of those with infection or malignancy (p < 0.001). As previously shown for anti-beta 2GPI antibodies, IgG2 was the predominant IgG subclass reactive with prothrombin. Thus, autoimmune patients with LA have a high incidence of antibodies to beta 2GPI and prothrombin, the binding of which could similarly require high antigen density and/or exposure of cryptic epitopes resulting from protein interaction with an irradiated (i.e. more anionic) polystyrene surface.
Arvieux
J
J
Laboratoire d'Immunologie, Centre de Transfusion Sanguine, Grenoble, France.
Darnige
L
L
Caron
C
C
Reber
G
G
Bensa
J C
JC
Colomb
M G
MG
D001323:Autoantibodies / Q000097:blood*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D016682:Lupus Coagulation Inhibitor / Q000097:blood / Q000276:immunology*; D011516:Prothrombin / Q000276:immunology*
D001323:Autoantibodies; D016682:Lupus Coagulation Inhibitor; D011516:Prothrombin