Selection of rabbit single-chain Fv fragments against the herbicide atrazine using a new phage display system

A convenient new bacteriophage display vector; pSD3, has been constructed a nd used to generate rabbit monoclonal anti-pesticide antibody fragments. Fo llowing amplification of immunoglobulin light chain, and heavy chain variab le region gene libraries, restriction enzymes Sfi I and PflM I are used to assemble scFv libraries in pSD3. This allows the number of stages involving the polymerase chain reaction and restriction enzyme digestion to be minim ized to optimize maintenance of the original diversity of the variable regi on genes in the libraries. The vector also incorporates an amber codon, a 6 xHis tag and a c-myc epitope to facilitate soluble single-chain Fv producti on detection and purification. Using the pSD3 system two anti-atrazine sing le-chain Fvs were,ere isolated from a library derived from the spleen cells of a rabbit immunized with bovine serum albumin-atrazine conjugate. Charac terization of single-chain Fvs by competition and equilibrium ELISA indicat ed good specificity and affinity, to atrazine.