Selection and characterisation of recombinant single-chain antibodies to the hapten Aflatoxin-B1 from naive recombinant antibody libraries.
Selection of antibodies from large repertoire phage display libraries has become a common technique for isolation of specific antibodies to antigens. Many of these libraries are shown to contain antibodies specific to haptens, but only when these haptens are derivatised or conjugated to an immobilising molecule, such as bovine serum albumin (BSA). There has been little demonstration of the suitability of naive recombinant antibody libraries for isolating antibodies that bind low molecular weight haptens in the absence of a carrier molecule and few have addressed the problems associated with selecting antibodies that only recognize the combination of hapten and the carrier molecule. We have panned two-phage antibody libraries against AflatoxinB1-BSA and screened single-chain antibody fragments for binding to AflatoxinB1-BSA and Aflatoxin-B1. Many of the antibodies isolated specifically bound AflatoxinB1-BSA, but not soluble Aflatoxin-B1 or BSA. Modification of the protocol led to isolation of single-chain fragment variable antibody domain (scFv) antibodies that specifically bound soluble Aflatoxin-B1 with an affinity of 6x10(-9) M.
Moghaddam
A
A
Affitech AS, Oslo Research Park, Gaustadalléen 21, N-0349 Oslo, Norway. amirmo@affitech.com
Løbersli
I
I
Gebhardt
K
K
Braunagel
M
M
Marvik
O J
OJ
D016604:Aflatoxin B1 / Q000737:chemistry / Q000276:immunology*; D000468:Alkalies; D000918:Antibody Specificity; D000941:Antigens / Q000276:immunology; D003429:Cross Reactions; D015870:Gene Expression; D006241:Haptens / Q000276:immunology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007128:Immunoglobulin Fragments / Q000235:genetics / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology*; D015394:Molecular Structure; D009183:Mycotoxins / Q000276:immunology; D019151:Peptide Library; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology*; D012995:Solubility; D020349:Surface Plasmon Resonance
D000468:Alkalies; D000941:Antigens; D006241:Haptens; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D009183:Mycotoxins; D019151:Peptide Library; D011994:Recombinant Proteins; C055966:immunoglobulin Fv; D016604:Aflatoxin B1