Immunochemical rapid test for multiresidue analysis of antimicrobial drugs in milk using monoclonal antibodies and hapten–glucose oxidase conjugates

Abstract

Based on monoclonal antibodies (Mabs) against streptomycin, sulfadiazine, sulfamethazine and cloxacillin, a semiquantitative immunochemical rapid test for multiresidue analysis has been developed. The test format is that of an indirect competitive enzyme immunoassay. Glucose oxidase conjugates of the individual compounds were found to be suitable coating antigens. Individual immunoassays exhibited sensitivities far below the respective maximum residue limits (MRLs) in milk, i.e. 3.2 ng ml−1 for streptomycin, 9.0 ng ml−1 for sulfadiazine, 7.0 ng ml−1 for sulfamethazine and 0.8 ng ml−1 for cloxacillin. By optimising the assay conditions, total assay time was reduced to 15 min. The individual tests were then integrated onto a single microtitre plate array, in which the milk samples were simultaneously incubated with a mixture of the specific Mabs. The detection limits obtained with the multitest system again were below the respective MRLs, i.e. 14, 20, 13 and 1.7 ng ml−1, respectively, for streptomycin, sulfadiazine, sulfamethazine and cloxacillin. The coefficients of variation were less than 10%. Cross-reactivities with structurally related substances have been determined for both the individual assays and the multitest system.