On the influence of vector design on antibody phage display

Abstract

Phage display technology is an established technology particularly useful for the generation of monoclonal antibodies (mAbs). The isolation of phagemid-encoded mAb fragments depends on several features of a phage preparation. The aims of this study were to optimize phage display vectors, and to ascertain if different virion features can be optimized independently of each other.Comparisons were made between phagemid virions assembled by g3p-deficient helper phage, Hyperphage, Ex-phage or Phaberge, or corresponding g3p-sufficient helper phage, M13K07. All g3p-deficient helper phage provided a similar level of antibody display, significantly higher than that of M13K07. Hyperphage packaged virions at least 100-fold more efficiently than did Ex-phage or Phaberge. Phaberge's packaging efficiency improved by using a SupE strain. Different phagemids were also compared. Removal of a 56 base pair fragment from the promoter region resulted in increased display level and increased virion production. This critical fragment encodes a lacZ'-like peptide and is also present in other commonly used phagemids.Increasing display level did not show statistical correlation with phage production, phage infectivity or bacterial growth rate. However, phage production was positively correlated to phage infectivity. In summary, this study demonstrates simultaneously optimization of multiple and independent features of importance for phage selection.

Progress in phage display: evolution of the technique and its applications

“…These helper phage can be amplified in amber suppressive host strains. A comparative study confirmed similar display levels of Fab fragments achieved by using hyperphage, ex-phage, or phaberge [82]. However, virion production and percentage of infective phagemid virions were highly dependent not only on the helper phage but also on the host strain and the phagemid used.…”

High-Affinity, Human Antibody-Like Antibody Fragment (Single-Chain Variable Fragment) Neutralizing the Lethal Factor (LF) of Bacillus anthracis by Inhibiting Protective Antigen-LF Complex Formation

“…The diversity of the library was estimated to be 1.13 ϫ 10 8 clones. The library was packaged using a protocol adapted from that for Fab (antigen binding fragment) phage production in reference 22: 2.5 ϫ 10 11 Hyperphage particles (19,48,53) were used. The packaging of the library was tested by titration and immunoblotting according to reference 22.…”

Semi-automated Magnetic Bead-Based Antibody Selection from Phage Display Libraries

“…For instance, biological parameters can be changed, i.e., different antibody libraries can be applied, or different helper phage systems can be used for initial packaging of the same library (e.g., Hyperphage versus M13K07; see Soltes et al 2007), or even different E. coli strains can be used for phage propagation. Furthermore, different buffer conditions or counter selection strategies can be introduced.…”