Background: The D antigen is a highly immunogenic human RBC antigen. Alloimmunization against the D antigen produces high-affinity antibodies that cause hemolytic transfusion reactions and HDN.
Study design and methods: Cloning and subsequent sequence analysis of 11 new samples of monoclonal anti-D was performed in an attempt to identify V(D)J germline gene usage. Sequences were compared and analyzed with 37 previously published samples of anti-D for identification of V(H) and V(L) pairings, canonical structures, and conformation of restricted germline gene usage.
Results: The V(H) and V(L) pairings used by the new D MoAbs resulted in seven canonical combinations, three of which had not been described previously. Preferential usage of gene segments from the VH3 and VH4 families and of D3, D6, JH6, and DPK9 germline gene segments was also determined. Three samples of anti-D from different donors were found to use similar V(H) and V(kappa) germline genes, despite the fact that two of the antibodies recognized epD6/7 and the third recognized epD1. From the cumulative analysis of the anti-D IgG, 24 V(H) and V(L) gene pairings were identified, resulting in only 10 canonical structures.
Conclusions: Despite the potential for diversity, only a minority of V(H) and V(L) germline genes are used by anti-D. Consequently, V(H) and V(L) pairings and the resulting canonical structures are similarly restricted.