title: Preparation of a Chicken scFv to Analyze Gentamicin Residue in Animal Derived Food Products.
abstract: Chicken is an ideal model for simplified recombinant antibody library generation. It has been rarely been reported to apply chicken single-chain variable fragments (scFvs) in immunoassays for the detection of antibiotic and chemical contaminants in animal food products. In this study, the scFvs (S-1 and S-5) were isolated from a phage display library derived from a hyperimmunized chicken. The checker board titration revealed that the optimum concentrations of S-1 and S-5 were 0.78 μg/mL and 0.44 μg/mL respectively, to obtain OD450 around 1.0 at 5 μg/mL of Gent-OVA coating concentration. Both S-1 and S-5 exhibited negligible cross reactivity with kanamycin and amikacin. The 50% inhibitory concentration (IC50) of S-1 and S-5 were 12.418 ng/mL and 14.674 ng/mL respectively. In the indirect competitive ELISA (ic-ELISA), the limits of detection for S-1 and S-5 were 0.147 ng/mL and 0.219 ng/mL respectively. The mean recovery for Gent ranged from 60.91% to 118.09% with no more than 10.35% relative standard deviation (RSD) between the intra-assay and the inter-assay. These results indicate the chicken scFv based ic-ELISA method is suitable for the detection of Gent residue in animal derived edible tissues and milk.
authors.affiliation: College of Veterinary Medicine, Northwest Agriculture and Forestry University , Yangling, 712100, Shaanxi, China.
authors.forename: Cui
authors.initials: C
authors.lastname: Li
authors.affiliation: College of Veterinary Medicine, Northwest Agriculture and Forestry University , Yangling, 712100, Shaanxi, China.
authors.forename: Jinxin
authors.initials: J
authors.lastname: He
authors.affiliation: College of Veterinary Medicine, Northwest Agriculture and Forestry University , Yangling, 712100, Shaanxi, China.
authors.forename: Hao
authors.initials: H
authors.lastname: Ren
authors.affiliation: College of Veterinary Medicine, Northwest Agriculture and Forestry University , Yangling, 712100, Shaanxi, China.
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Zhang
authors.affiliation: College of Veterinary Medicine, Northwest Agriculture and Forestry University , Yangling, 712100, Shaanxi, China.
authors.forename: Enqi
authors.initials: E
authors.lastname: Du
authors.affiliation: College of Veterinary Medicine, Northwest Agriculture and Forestry University , Yangling, 712100, Shaanxi, China.
authors.forename: Xinping
authors.initials: X
authors.lastname: Li
mesh_terms: D000818:Animals; D002645:Chickens / Q000276:immunology*; D004350:Drug Residues / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay; D005506:Food Contamination / Q000032:analysis*; D005839:Gentamicins / Q000032:analysis* / Q000276:immunology*; D057127:Single-Chain Antibodies / Q000276:immunology* / Q000302:isolation & purification
chemical_list: D005839:Gentamicins; D057127:Single-Chain Antibodies