Immunization of a rabbit with β 2-glycoprotein I induces charge-dependent crossreactive antibodies that bind anionic phospholipids and have similar reactivity as autoimmune anti-phospholipid antibodies

Abstract

A rabbit immunized with β 2-glycoprotein I (β 2-GPI) produced Abs that bind to negatively charged phospholipids and to β 2-GPI. After affinity purification of the Abs to β 2-GPI, the dual reactivity could still be detected. Adsorption studies with a phosphatidylserine affinity column depleted phospholipid-reactive Abs, but β 2-GPI reactivity was retained. The same pattern of reactivity was found with culture supernatants from rabbit anti-β 2-GPI splenocytes fused with an immortalized rabbit cell line. The reactivity to negatively charged phospholipids is likely to involve ionic interactions, as high ionic strength buffers eliminated binding to anionic phospholipids, but not to β 2-GPI. Affinity-purified anti-phospholipid (aPL) Abs from four of seven autoimmune patients bound anionic phospholipids in the absence of β 2-GPI. However, in high ionic strength buffer, this binding was abolished in three patients and significantly reduced in the fourth. In contrast, affinity-purified aPL Abs from seven autoimmune patients bound to β 2-GPI-coated plates, and binding in high ionic strength buffer was reduced only moderately in three patients. Therefore, autoimmune-type aPL Abs display anti-β 2-GPI reactivity and charge-dependent binding to anionic phospholipids similar to affinity-purified rabbit anti-β 2-GPI Abs.