Display of binding agents
The present invention relates to a method of preparing a genetic package displaying oligomers of modular antibody domains binding to a target and to a scaffold ligand as well as to vectors and libraries of genetic packages produced thereby. The invention further relates to methods of selecting suitable linker sequences for use in such oligomer display.
1. A method of producing a functional binding agent, wherein the functional binding agent binds to a target antigen, wherein the functional binding agent is an antibody Fc fragment consisting of a CH2 domain and a CH3 domain, comprising the steps of:
a) providing a library of said antibody Fc fragments,
b) contacting said library with said target antigen in the presence of a scaffold ligand, wherein said target antigen and said scaffold ligand are different molecules, wherein said scaffold ligand is selected from the group consisting of CD64, CD16, CD32, FcRn and Protein A,
c) selecting a library member binding to said target antigen in the presence of said scaffold ligand to obtain a functional binding agent, and
d) producing a preparation of the functional binding agent.
2. The method of claim 1 , wherein said library contains at least 10 2 independent clones expressing functional binding agents.
3. The method of claim 1 , wherein the functional binding agent has a target binding affinity of Kd<10 −8 M.
4. The method of claim 1 , wherein said target antigen is a receptor of the erbB class.
5. The method of claim 1 , further comprising the steps of:
e) affinity maturating the functional binding agent by amino acid variation to obtain an affinity matured pool of binding agents, and
f) selecting a member of said pool which binds to the target antigen in the presence of the scaffold ligand to obtain an affinity maturated functional binding agent.
6. The method of claim 5 , wherein the affinity maturated functional binding agent exhibits at least a 10 fold increase in affinity of binding to the target antigen compared to the functional binding agent.