Selection of binders from phage displayed antibody libraries using the BIAcore biosensor.
In this report we show that phage displayed antibodies can be selected based on dissociation rate constants, using a BIAcore biosensor. To demonstrate the principle, two Fab phage stocks displaying antibodies specific for hen egg lysozyme or phenyloxazolone were mixed in a ratio of 1:10 and injected over the biosensor chip containing immobilized lysozyme. Antigen-specific bound phages were eluted and analysed for specificity and phage titer. This procedure enriched for phages carrying specific antibodies. Selection of high affinity binders from phage libraries was then demonstrated with the BIAcore when phages were eluted and collected at different time points. Soluble antibody fragments were subsequently expressed and their kinetic parameters were determined. The time of elution was directly proportional to the affinity, due to decreased dissociation rate constants. This procedure offers a rapid and simple approach for selecting binders from phage libraries differing in antibody dissociation rate constants.
Department of Immunotechnology, Lund University, Sweden.
A C
AC
Malmborg
M
M
Dueñas
M
M
Ohlin
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E
Söderlind
C A
CA
Borrebaeck
D001435:Bacteriophages / Q000378:metabolism*; D001483:Base Sequence / Q000235:genetics; D001666:Binding Sites, Antibody / Q000276:immunology*; D015374:Biosensing Techniques / Q000145:classification*; D002999:Clone Cells; D015723:Gene Library; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000276:immunology; D007700:Kinetics; D009113:Muramidase / Q000276:immunology; D011485:Protein Binding / Q000276:immunology
D007140:Immunoglobulin Fab Fragments; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D009113:Muramidase