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Genetic and Immunological Properties of Phage-Displayed Human
Anti-Rh(D) Antibodies: Implications for Rh(D) Epitope Topology
Tylis Y. Chang and
Don L. Siegel
From the Blood Bank/Transfusion Medicine Section, Department of
Pathology & Laboratory Medicine, University of Pennsylvania School of
Medicine, Philadelphia, PA.
Understanding anti-Rh(D) antibodies on a molecular level would
facilitate the genetic analysis of the human immune response to Rh(D),
lead to the design of therapeutically useful reagents that modulate
antibody binding, and provide relevant information regarding the
structural organization of Rh(D) epitopes. Previously, we described a
Fab/phage display-based method for producing a large array of
anti-Rh(D) antibodies from the peripheral blood lymphocytes of a single
alloimmunized donor. In the current study, we present a detailed
analysis of 83 randomly selected clones. Sequence analysis showed the
presence of 28 unique  1 heavy chain and 41 unique light
chain gene segments. These paired to produce 53 unique Fabs that had
specificity for at least half of the major Rh(D) epitopes.
Surprisingly, despite this diversity, only 4 closely related heavy
chain germline genes were used (VH3-30, VH3-30.3, VH3-33, and VH3-21).
Similarly, nearly all V light chains (15/18) were
derived from one germline gene (DPK9).  light chains showed a more
diverse VL gene usage, but all (23/23) used the identical
J 2 gene. Several Fabs that differed in epitope specificity used identical heavy chains but different light chains. In
particular, 2 such clones differed by only 3 residues, which resulted
in a change from epD2 to epD3 specificity. These results suggest a
model in which footprints of anti-Rh(D) antibodies are essentially
identical to one another, and Rh(D) epitopes, as classically defined by
panels of Rh(D) variant cells, are not discrete entities. Furthermore,
these data imply that the epitope specificity of an anti-Rh(D) antibody
can change during the course of somatic mutation. From a clinical
perspective, this process, which we term epitope migration, has
significance for the design of agents that modulate antibody production
and for the creation of mimetics that block antibody binding in the
settings of transfusion reactions and hemolytic disease of the newborn.
Blood, Vol. 91 No. 8 (April 15), 1998:
pp. 3066-3078
© 1998 by The American Society of Hematology.
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