Development of a biotin-avidin mediated immunoassay for simultaneous detection of danofloxacin, enrofloxacin and ciprofloxacin in milk

摘要

To generate a group-specific monoclonal antibody (McAb) against danofloxacin (DANO), enrofloxacin (ENR) and ciprofloxacin (CIP), glutaraldehyde was used to link the presentative hapten of CIP to the immunogen and coating antigen, respectively, leaving the common structure of these 3 drugs exposed in both conjugates as a major antigenic site. Consequently, a McAb (6D3) with high cross-reactivity to these three antibiotics has been obtained by using hybridomas technique. In a biotin-avidin mediated enzyme-linked immunosorbent assay, the IC50 values for DANO, ENR and CIP were 5.1, 4.5 and 4.2 ng mL~(-1), respectively. The ELISA was used for the detection of spiked DANO and ENR+CIP in milk. The recoveries ranged from 74.1 to 92.4% and coefficients of variation were in a range of 6.6-11.9%. The accuracies and sensitivity of the method were good for simultaneous analysis of the 3 drugs in milk after a simple sample extraction process.