The potential of monoclonal antibodies against ampicillin for the preparation of a multi-immunoaffinity chromatography for penicillins.
Monoclonal antibodies (Mab) against ampicillin were prepared by immunization of mice with an ampicillin-keyhole limpet hemocyanin conjugate coupled by a glutaraldehyde method. Sensitivity and specificity of these antibodies were tested in a direct competitive enzyme immunoassay, in which an ampicillin-horseradish peroxidase conjugate prepared by a carbodiimide method served as the labelled antigen. According to their cross-reactivities with the other beta-lactam antibiotics, the Mabs could be divided into two groups, which are represented by the clones designated 1D1 and 3B5. While Mab 3B5 (IgG1) showed no major cross-reactions with the other penicillins frequently used in veterinary medicine except for amoxicillin (108%), Mab 1D1 (IgG2a) had marked cross-reactivities with most of the 17 tested beta-lactam antibiotics (e.g., amoxicillin 187%, penicillin G 31%, cloxacillin 30%, dicloxacillin 44%, and oxacillin 14%). The detection limits for ampicillin, calculated from the antibiotic concentration giving 30% binding inhibition, were 11.7 (Mab 3B5) and 16.6 ng ml-1 (Mab 1D1). To prepare multi-immunoaffinity chromatography columns, Mab 1D1 and a previously described antibody against cloxacillin (Mab 1F7) were each coupled to CNBr activated sepharose. The capacity of the resulting immunosorbents was approximately 6.6 and 5.4 micrograms ml-1 gel for ampicillin and cloxacillin, respectively. Recoveries of amoxicillin, ampicillin, cloxacillin, dicloxacillin, penicillin G and oxacillin (in buffer solutions) from the produced immunoaffinity columns were in the range from 67 to 100%.
Dietrich
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R
Institute for Hygiene and Technology of Food of Animal Origin, Veterinary Faculty, University of Munich, Germany.
Usleber
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E
Märtlbauer
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D000667:Ampicillin / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification; D002846:Chromatography, Affinity / Q000379:methods; D004350:Drug Residues / Q000032:analysis*; D005260:Female; D005506:Food Contamination / Q000032:analysis*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008815:Mice, Inbred Strains; D010406:Penicillins / Q000032:analysis* / Q000737:chemistry; D019155:Veterinary Drugs / Q000032:analysis*
D000911:Antibodies, Monoclonal; D010406:Penicillins; D019155:Veterinary Drugs; D000667:Ampicillin