Production of high affinity monoclonal antibody and development of indirect competitive chemiluminescence enzyme immunoassay for gentamicin residue in animal tissues

Abstract

To determine gentamicin residues in animal tissues, a monoclonal antibody (Mab) was produced and a sensitive indirect competitive chemiluminescent enzyme immunoassay (icCLEIA) was developed. At first, gentamicin was conjugated with bovine serum albumin as immunogens which were used to immunize BALB/c mice. Then, an anti-gentamicin Mab was prepared by hybridoma technology. Finally, a sensitive icCLEIA was developed with an 50% inhibition concentration (IC50) of 0.067 ng/mL for gentamicin. The limit of detection of the icCLEIA was 0.002 ng/mL. The cross reactivity of the Mab with structural analogues were<0.01%. The recoveries of gentamicin ranged from 80 to 101% and coefficient of variation was <6.4% in pork and fish samples. Samples were detected by UPLC-MS/MS for evaluating reliability of the icCLEIA. The results suggested that the prepared anti-gentamicin Mab can be used for rapid and convenient immunoassays to detect gentamicin residues in animal tissues.