Synthesis of Rh Fv phage-antibodies using VH and VL germline genes
Nevin Hughes-Jones
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorJacqueline Bye
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorBarbara Gorick
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorJames Marks
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorWillem Ouwehand
Department of Haematology, University of Cambridge, Cambridge, and National Blood Service East Anglia
Search for more papers by this authorNevin Hughes-Jones
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorJacqueline Bye
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorBarbara Gorick
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorJames Marks
Molecular Immunopathology Unit and Centre for Protein Engineering, Medical Research Council, Cambridge,
Search for more papers by this authorWillem Ouwehand
Department of Haematology, University of Cambridge, Cambridge, and National Blood Service East Anglia
Search for more papers by this authorAbstract
Antibodies to the D antigen of the Rh system use a restricted set of immunoglobulin V and J gene segments, especially VH DP50 and DP63, JH6, Vλ DPL16 and Jλ 2/3. These gene segments may confer a natural affinity on the antibodies for the D antigen and this hypothesis has been tested by constructing two single-chain Fv phage-antibody libraries based on the germline gene segments DP50 and DP63; structural variability was obtained by insertion of 11 amino acids in random sequence in the VHCDR3. 10 anti-D antibodies were selected from these libraries, each with a unique VHCDR3. In contrast, selections with the CcEe antigens produced antibodies reacting with the Rh polypeptide molecules but without strict blood group specificity. One of these latter DP50-based antibodies was converted into 12 different antibodies with specificity for E by replacing the original germline light chain with chains from a rearranged L chain library. The CDR1 and CDR2 sequences of the DP50-based antibodies were common to both anti-D and anti-E molecules; differentiation between D and E specificity was dependent on VHCDR3 sequences and their correct pairing with an appropriate L chain.
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