Antibody library selection by the {beta}-lactamase protein fragment complementation assay.
Protein fragment complementation assay (PCA) is based on the interaction between two protein partners (e.g. target antigen and antibody), which are genetically fused to the two halves of a dissected marker protein. Binding of the two partners reassembles the marker protein and hence reconstitutes its activity. In this work we have developed the first application of beta-lactamase-based PCA for the isolation of single chain Fv fragments (scFvs) binding to the human receptor RON from a naïve library. Specific scFvs with the ability to immunoprecipitate could be isolated after a single round of PCA selection from an scFv repertoire previously pre-selected by phage display. Furthermore, the PCA was used to successfully map the epitopes recognized by the selected scFvs by screening them against a small library of random RON fragments.
Secco
Paola
P
Department of Medical Sciences, Interdisciplinary Research Centre on Autoimmune Diseases, University of Eastern Piedmont, Novara.
D'Agostini
Elena
E
Marzari
Roberto
R
Licciulli
Marta
M
Di Niro
Roberto
R
D'Angelo
Sara
S
Bradbury
Andrew R M
AR
Dianzani
Umberto
U
Santoro
Claudio
C
Sblattero
Daniele
D
D000818:Animals; D000906:Antibodies / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D000915:Antibody Affinity / Q000235:genetics; D000941:Antigens / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D018604:Epitope Mapping; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D051379:Mice; D010446:Peptide Fragments / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D019151:Peptide Library; D040081:Protein Array Analysis / Q000379:methods*; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism
D000906:Antibodies; D000941:Antigens; D010446:Peptide Fragments; D019151:Peptide Library; D011993:Recombinant Fusion Proteins; C080835:RON protein; D020794:Receptor Protein-Tyrosine Kinases; D001618:beta-Lactamases