Selection of VHHs under application conditions.

The successful application of antibody fragments such as VHHs in diagnostic assays, affinity purification, imaging, or therapy is not determined by the specificity and affinity of the antibody fragment alone. The ability to bind the target protein in the environment in which the antibody fragment is intended to functionally perform determines to a great extent its success. To identify antibodies with the required stability profile selection of naturally occurring variants from an immune library or mutants from an engineered library should be performed via phage display. The conditions under which the designed antibodies displayed on phage bind to the target ideally should mimic the environment in which the antibody should be effective. After selection individual antibodies need to be tested in appropriate screening assays, again taking into account the conditions under which the antibody should bind to the target and induce the desired effect.