Expression of antibody Fab domains on bacteriophage surfaces. Potential use for antibody selection.
We describe a method for expressing an antibody Fab fragment on the surface of M13 filamentous bacteriophage. The L chain gene, preceded by an Escherichia coli signal sequence, is linked to the 5' end of the gene for the M13 major coat protein. A partial H chain gene, preceded by an E. coli signal sequence and truncated at the end of the CH1 region, is inserted on a plasmid adjacent to the fused L chain gene, so both genes are under the transcriptional control of an inducible promoter. The plasmid also contains the M13 origin of replication. When the promoter is induced, functional antibody Fab fragment appears on the surface of the E. coli inner membrane, as shown by specific binding to an Ag-affinity matrix. When the E. coli are further infected with a helper phage, allowing replication and packaging of the plasmid into phage particles, the resulting phage specifically bind to an Ag-coated plate, indicating they have antibody Fab fragment on their surface. The antibody-expressing phage can also be specifically bound to and eluted from an Ag-affinity column. These observations support the possibility of a new way of generating antibodies, in which amplified Ig cDNA from an appropriate B cell population is cloned into a suitable M13 vector, and phage containing the genes for desired antibodies are selected directly by binding to an Ag-affinity matrix.
Protein Design Labs, Inc., Mountain View, CA 94043.
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Chang
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Landolfi
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C
Queen
D002846:Chromatography, Affinity; D003090:Coliphages / Q000235:genetics*; D004926:Escherichia coli / Q000276:immunology; D005822:Genetic Vectors; D007140:Immunoglobulin Fab Fragments / Q000235:genetics*; D008565:Membrane Proteins / Q000276:immunology; D010957:Plasmids; D016133:Polymerase Chain Reaction; D015375:Receptors, Interleukin-2 / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology
D007140:Immunoglobulin Fab Fragments; D008565:Membrane Proteins; D015375:Receptors, Interleukin-2; D011994:Recombinant Proteins