Use of Escherichia coli Mutator Cells to Mature Antibodies

Despite the power of antibody (Ab) phage-display technology, a problem which can be commonly encountered is the recovery of Abs of low affinity for the antigen (Ag) of interest. Two general strategies can be applied to increase affinity: mutations can be scattered randomly throughout the genes; substitutions can be introduced in a directed manner to specific regions, such as the complementarity-determining loops. In order to select those changes that improve on the starting affinity for the target Ag, phage display can be utilized, the power of this approach lying in the display of Abs at the viral surface coupled with carriage of the encoding sequences within the phage particle. Repeated rounds of mutation and increasingly stringent selection (Fig. 1 ) enable recovery of Abs of substantially elevated affinity for the target. In general, the greatest improvements in affinity are observed when low-affinity Abs
(Kd
<10−
6M)
are used as the starting point. Although high-affinity Abs
(Kd>
10−
8
M) are less readily improved, there have been isolated successes.