Direct binding of lupus anticoagulant antibodies to nonbilayer phosphatidylethanolamine

The present study describes a new phase-specific assay system for the detec tion of anti-phospholipid antibodies, based on the finding by P-31 NMR anal ysis that phospholipid can be quantitatively retained on nitrocellulose pap er in a phase-sensitive fashion. Using this: system, we demonstrate that hu man hybridoma lupus anticoagulant antibodies bind directly to nonbilayer ph ase phosphatidylethanolamine, a lipid architecture that we have shown speci fically inhibits lupus anticoagulant activity and is recommended for confir mation of the diagnosis of these antibodies. We have analyzed 33 human hybr idoma antibodies, of which 16 had lupus anticoagulant antibody activity. Se venty-five percent of the lupus anticoagulant antibodies bound directly to nonbilayer phase phosphatidylethanolamine, while only 12% bound to immobili zed lamellar phase phosphatidylethanolamine. In contrast, none of the 17 hy bridoma antibodies without lupus anticoagulant activity bound to either lam ellar or nonbilayer phase phosphatidylethanolamine. Forty-four percent and 62%, respectively, of the lupus anticoagulant antibodies bound to dioleoylp hosphatidylserine and cardiolipin, both negatively charged bilayer phase ph ospholipids. These data provide the first direct demonstration of the prefe rential reactivity of human lupus anticoagulant antibodies with nonbilayer phosphatidylethanolamine. The structurally sensitive solid phase assay syst em described here provides the means to study a variety of phospholipid epi topes and to further analyse the role of phospholipid architecture in anti- phospholipid anti-body syndromes.