Development of an indirect competitive ELISA for detection of danofloxacin residue in milk

In order to develop the immunoassay method of detecting residue of Danofloxacin (DFLX) in milk, DFLX was used as a hapten which was coupled with Bovine serum albumin (BSA) and DFLX-BSA was formed. Then the rabbits were immunized with it and the specificity antibody against DFLX was obtained. The assay performance was improved by use of a coating antigen (DFLX-OVA), which was a conjugation of the DFLX and Ovalbumin (OVA). Finally, the determination of DFLX residue by Indirect Competitive ELISA (Ci-ELISA) was studied. The results of the experiment demonstrated that limit of detection (LOD) for DFLX was 29.24 μg/kg. Besides, the specificity antibody crossly reacted with the closely related FQNs, including Ciprofloxacin (CPLX), Ofloxacin (OFLX), Sarafloxacin (SALX) and Enrofloxacin (EFLX). The average recovery rate of standard substance (DFLX) is 91.45% within the range of 0.5~ 2 000 μg/kg in milk. The developed method was practicable to screen the residues of DFLX in milk.