Multimerization domains for antibody phage display and antibody production.
High-throughput generation of antibodies for proteome research has become feasible by using antibody gene libraries and in vitro selection methods like phage display. Typically monovalent antibody fragments like scFv, Fab or scFab are obtained by this technology. To mimic the IgG molecule and gain avidity, resulting in stronger binding, multimerization domains can be fused to antibody fragments. Here we systematically analyzed different multimerization domains in respect to three key parameters, crucial for the high-throughput generation of binders. (i) The compatibility to be displayed on phage (assessed for at least three different antibody formats, scFv, Fab and scFab) in combination with five different multimerization domains; (ii) production yields and (iii) oligomerization properties were analyzed for three different scFv fragments. We found that the use of a biotin acceptor domain in combination with an in vivo biotinylation system performed best concerning the key parameters and thus would be a useful tool to generate multimeric antibody complexes on demand from phage display selected antibody fragments with the least effort.
Thie
H
H
Technische Universität Braunschweig, Institut für Biochemie und Biotechnologie, Abteilung Biotechnologie, Spielmannstr. 7, 38106 Braunschweig, Germany. h.thie@tu-bs.de
Binius
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S
Schirrmann
T
T
Hust
M
M
Dübel
S
S
D000906:Antibodies / Q000737:chemistry* / Q000276:immunology*; D000917:Antibody Formation / Q000276:immunology*; D000941:Antigens / Q000276:immunology; D004926:Escherichia coli; D007135:Immunoglobulin Variable Region / Q000276:immunology; D010446:Peptide Fragments / Q000276:immunology; D019151:Peptide Library; D011485:Protein Binding; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D012995:Solubility; D019809:Streptavidin / Q000276:immunology
D000906:Antibodies; D000941:Antigens; D007135:Immunoglobulin Variable Region; D010446:Peptide Fragments; D019151:Peptide Library; D019809:Streptavidin