Characterization of monoclonal anti-β2-glycoprotein-I and anti-prothrombin antibody fragments generated by phage display from a patient with primary antiphospholipid syndrome

The molecular structure of antibodies associated with autoimmune thrombosis is beginning to be understood. We describe the binding specificities and sequence analysis of anti-β2-glycoprotein-I (anti-β2GP-I) or anti-prothrombin (anti-PT) antibody fragments generated by phage display from a patient with primary antiphospholipid syndrome (APS). We obtained 39 positive clones, two that had the correct size reacted with β2GP-I (Beta 1 and Beta 2). Ten clones with the same restrictive pattern recognized PT (Prot 1) and cross-reacted with β2GP-I. All three clones recognized anionic and zwitterionic phospholipids. The VH regions of both anti-β2GP-I clones are members of the VH4 family. Prot 1 has a VH segment of the VH3 family. The Beta 1 JH segments are JH5b and JH4b for Beta 2 and Prot 1. VL genes are Vλ1, 3 and 1, respectively. No JL was identified for Beta 1, while Beta 2 and Prot 1 carry Jλ3b genes. Beta 1 and Beta 2 carry highly conserved germ-line VH and VL genes. Mutations of the Prot 1 gene appear to be antigen-dependent, most are hotspot mutations located in the CDR 1 and 2 regions. Our work suggests that some anti-β2GP-I from patients with primary APS are natural autoantibodies. Our work may also help to explain the frequent coexistence of anti-β2GP-I and anti-PT in the same patient.