MOLECULAR-CLONING OF MURINE MONOCLONAL ANTIIDIOTYPIC FAB

Anti-idiotypic antibodies (Ab2) binding to idiotopes on antibodies with various antigen binding specificities (Ab1) are potential regulators of immunity in a variety of diseases, such as autoimmunity, cancer, and viral, bacterial, or parasitic infections. Furthermore, Ab2 are useful probes for the characterization of receptor/ligand interactions. Thus far, Ab2 production has been limited to the isolation of polyclonal Ab2 from immune sera or monoclonal Ab2 from hybridoma supernatants. However, both approaches have produced a limited number of Ab2. As an alternative approach, we demonstrate here the production of Ab2-Fab by using repertoire cloning. Using HIV-1 as a model system, the Ab2-Fab were generated from the spleens of mice immunized with the virus-neutralizing and syncytia-inhibiting anti-HIV-1 monoclonal antibody 0.5beta. A bacteriophage lambda vector system was used to express a combinatorial library in Escherichia coli. Iodinated 0.5beta was used to identify 17 Ab2-Fab clones. DNA sequence analysis of five clones revealed three similar kappa and Fd combinations. The Ab2-Fab bound with high affinity (3.5-6.5 x 10(9) liters/mol) specifically to the Ab1 and not to isotype-matched antibodies with unrelated specificities. The three Ab2-Fab probably bind to the same idiotope on the Ab1 as demonstrated in cross-competition binding studies. The Ab2-Fab inhibited binding of the Ab1 to antigen, and therefore, may functionally mimic the epitope defined by the Ab1 Repertoire cloning of Ab2-Fab may facilitate the generation of-Ab2 that have potential as modulators of immune responses against various antigens.