Production and characterization of monoclonal and recombinant antibodies against antimicrobial sulfamethazine.
A monoclonal antibody (mab) against the antimicrobial sulfamethazine was prepared and characterized by an indirect competitive enzyme-linked immunosorbent assay (IC-ELISA). Sulfamethazine in the range of 0.2 and 45 ng/ml could be determined with the mab by IC-ELISA. cDNAs encoding a variable heavy chain and variable light chain of the mab were cloned to produce recombinant antibodies using phage display technology. Following phage rescue and three rounds of panning, a single-chain variable fragment (scFv) antibody with high sulfamethazine-binding affinity was obtained. ELISA analysis revealed that scFv antibody and parent mab showed similar, but not identical, characteristics. The IC50 value by IC-ELISA with scFv antibody was 4.8 ng/ml, compared with 1.6 ng/ml with the parent mab. Performances of the assays in the presence of milk matrix were compared; the mab-based assay was less affected than the scFv-based assay. Sixty milk samples were analyzed by mab-based IC-ELISA, and four samples were sulfamethazine positive; these results were favorably correlated with those obtained by HPLC.
Yang
Zheng-You
ZY
Division of Applied Life Science, Graduate School of Gyeongsang National University, Jinju 660-701, Korea.
Shim
Won-Bo
WB
Kim
Min-Gon
MG
Lee
Kyu-Ho
KH
Kim
Keun-Sung
KS
Kim
Kwang-Yup
KY
Kim
Cheol-Ho
CH
Ha
Sang-Do
SD
Chung
Duck-Hwa
DH
D000595:Amino Acid Sequence; D000818:Animals; D000890:Anti-Infective Agents / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D001483:Base Sequence; D002417:Cattle; D002459:Cell Fusion; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D007128:Immunoglobulin Fragments / Q000096:biosynthesis; D051379:Mice; D008807:Mice, Inbred BALB C; D008892:Milk / Q000737:chemistry; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis*; D013418:Sulfamethazine / Q000032:analysis / Q000276:immunology*
D000890:Anti-Infective Agents; D000911:Antibodies, Monoclonal; D007128:Immunoglobulin Fragments; D011994:Recombinant Proteins; C055966:immunoglobulin Fv; D013418:Sulfamethazine