Abstract: The present invention relates to a method for in vivo molecular evolution of antibody function. According to the present invention, a nucleic acid encoding a CDR that is normally contained in a framework (the “original framework”), which differs from a selected master framework, is amplified from an immunoglobulin gene and is inserted into a nucleic acid encoding the selected master framework. The invention further provides an antibody library, such as a phage display library, and methods of making the same.

Abstract: The invention relates to methods of making anti-ligand libraries, which anti-ligands have a cavity binding site. The method comprises providing a plurality of anti-ligands having a cavity for binding a ligand, said plurality of anti-ligands being derived from a first anti-ligand having a cavity for binding a first ligand and differing from the first anti-ligand in that one or more of the amino acid residues which make up the ligand binding surface of the cavity are varied whereby the plurality of anti-ligands have a binding affinity for a first ligand which is different from that of the first anti-ligand, and wherein the one or more amino acid residues which is varied, is varied by replacement with one amino acid having similar but different physicochemical properties to the one or more amino acid residues, and wherein the one or more amino acids being varied can be varied to become a different amino acid in each anti-ligand molecule being varied.

Abstract: The present invention provides a method for producing a polynucleotide sequence encoding an antibody variable domain, the variable domain comprising complementarity-determining regions (CDRs) located within a selected framework (the ‘master framework’), the method comprising the steps of (a) providing at least one nucleic acid molecule encoding one or more CDRs and associated framework regions (the ‘original framework’), (b) amplifying at least one CDR-encoding portion of the nucleic acid molecule(s) of step (a) using one or more pairs of oligonucleotides as amplification primers and (c) assembling a polynucleotide sequence encoding an antibody variable domain by combining the amplified CDR-encoding nucleotide sequences produced in step (b) with nucleotide sequences encoding said master framework, wherein the oligonucleotide primers of step (b) comprise nucleotide sequences which differ from the corresponding nucleotide sequences encoding said master framework.