Antibody fragments from a 'single pot' phage display library as immunochemical reagents.
The display of repertoires of antibody fragments on the surface of filamentous bacteriophage offers a new way of making antibodies with predefined binding specificities. Here we explored the use of this technology to make immunochemical reagents to a range of antigens by selection from a repertoire of > 10(8) clones made in vitro from human V gene segments. From the same 'single pot' repertoire, phage were isolated with binding activities to each of 18 antigens, including the intracellular proteins p53, elongation factor EF-1 alpha, immunoglobulin binding protein, rhombotin-2 oncogene protein and sex determining region Y protein. Both phage and scFv fragments secreted from infected bacteria were used as monoclonal and polyclonal reagents in Western blots. Furthermore the monoclonal reagents were used for epitope mapping (a new epitope of p53 was identified) and for staining of cells. This shows that antibody reagents for research can be readily derived from 'single pot' phage display libraries.
Nissim
A
A
MRC Centre for Protein Engineering, Cambridge, UK.
Hoogenboom
H R
HR
Tomlinson
I M
IM
Flynn
G
G
Midgley
C
C
Lane
D
D
Winter
G
G
D000595:Amino Acid Sequence; D000818:Animals; D000918:Antibody Specificity; D001435:Bacteriophages; D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D003001:Cloning, Molecular; D017931:DNA Primers; D000939:Epitopes; D015723:Gene Library; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics; D007158:Immunologic Techniques; D007202:Indicators and Reagents; D008969:Molecular Sequence Data
D017931:DNA Primers; D000939:Epitopes; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D007202:Indicators and Reagents