title: Development of a class-specific polyclonal antibody-based indirect competitive ELISA for detecting fluoroquinolone residues in milk.
abstract: Modified 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) method was employed to synthesize the artificial antigen of norfloxacin (NOR), and New Zealand rabbits were used to produce anti-NOR polyclonal antibody (pAb). Based on the checkerboard titration, an indirect competitive enzyme-linked immunosorbent assay (icELISA) standard curve was established. This assay was sensitive and had a working range from 0.12 to 68.40 ng/ml, with the half maximal inhibitory concentration (IC(50)) and limit of detection (LOD) values of 2.7 ng/ml and 0.06 ng/ml, respectively. The produced pAb exhibited high cross-reactivity to fluoroquinolones (FQs) tested, and the IC(50) values to enoxacin, ciprofloxacin, and pefloxacin were 3.1, 3.4, and 4.1 ng/ml, respectively. It also indicated that the concentrations of NaOH and methanol in assay buffer should not be higher than 10% and 30%. When spiked in milk at 5, 20, and 50 ng/ml, the recoveries for NOR, enoxacin, ciprofloxacin, and pefloxacin ranged 90.5%-98.0%, 84.0%-95.2%, 94.0%-106.0%, and 89.5%-100.0%, respectively. The results suggest that this class-specific pAb-based icELISA could be utilized for the primary screening of FQ residues in animal-original products.
authors: College of Veterinary Medicine, Northwest A&F University, Yangling, China.
authors: Guo-ying
authors: GY
authors: Fan
authors: Ruo-song
authors: RS
authors: Yang
authors: Jin-qing
authors: JQ
authors: Jiang
authors: Xin-yao
authors: XY
authors: Chang
authors: Jun-jie
authors: JJ
authors: Chen
authors: Yong-hua
authors: YH
authors: Qi
authors: Shi-xiu
authors: SX
authors: Wu
authors: Xue-feng
authors: XF
authors: Yang
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology; D002417:Cattle; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D024841:Fluoroquinolones / Q000032:analysis* / Q000276:immunology; D005504:Food Analysis / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D008892:Milk / Q000737:chemistry* / Q000276:immunology; D015202:Protein Engineering / Q000379:methods; D011817:Rabbits; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: C429108:107-1A4 monoclonal antibody; D000911:Antibodies, Monoclonal; D024841:Fluoroquinolones