High affinity ScFvs from a single rabbit immunized with multiple haptens.
We report the generation of single-chain Fv (scFv) fragments with high affinities against four different hapten molecules from a single immunised rabbit. The rabbit was immunised with a mixture of protein conjugates of four different haptens, namely the herbicide mecoprop and derivatives of the herbicides atrazine, simazine, and isoproturon. An scFv phage display library was constructed, and several scFvs with high affinity against each hapten were isolated. For each hapten, a single binder was selected by k(off) ranking and used for affinity determination. The affinities were in sub-nanomolar range and the lowest K(d) value obtained was 6.75 x 10(-10) M. An unusual feature of one of the anti-isoproturon scFvs was its ability to retain binding activity at pH1.7. The utility and potential of using a single animal and immunisation with multiple antigens for the production of multiple, specific, high affinity scFvs by phage display is discussed.
Li
Y
Y
Department of Biology, University of Leicester, University Road, Leicester, LE1 7RH, United Kingdom. yl4@le.ac.uk
Cockburn
W
W
Kilpatrick
J B
JB
Whitelam
G C
GC
D000595:Amino Acid Sequence; D000818:Animals; D005260:Female; D006241:Haptens / Q000276:immunology*; D007114:Immunization; D007128:Immunoglobulin Fragments / Q000276:immunology*; D008969:Molecular Sequence Data; D011817:Rabbits; D017386:Sequence Homology, Amino Acid
D006241:Haptens; D007128:Immunoglobulin Fragments