Entwicklung eines Biosensors zum Nachweis von Antibiotika und Sulfonamiden in Milch- Herstellung der immunchemischen Komponenten

This paper describes the development and application of enzyme immunoassays for the detection of antimicrobials in milk, aiming at the establishment of a biosensor system for the on-line analysis of drug residues in milk. For the development of group-specific antibodies against penicillins rabbits were immunized with an ampicillin-BSA-conjugate. The resulting antiserum was employed for the development of a direct competitive enzyme immunoassay (EIA), giving a detection limit of 1 ng/ml for penicillin G in milk. Due to broad cross-reactivities the sensitive detection of those penicillins regulated by MRLs within the European Union (ampicillin, amoxicillin, oxacillin, cloxacillin, dicloxacillin and nafcillin) was enabled. The practical use of the enzyme immunoassay was demonstrated by analyzing artificially contaminated and violative incurred milk samples (n = 321). For the development of indirect competitive enzyme immunoassays for the detection of streptomycin, sulfonamides and penicillins, previously established direct assays, based on monoclonal antibodies, were adapted to indirect formats. For this purpose a wide range of coating-antigens was prepared by linking haptens to carrier-proteins. After optimizing test sensitivity and characterizing the test specificity indirect EIAs could be developed for each antimicrobial compound, fulfilling the MRL requirements due to EU regulation 2377/90. Only for ampicillin and penicillin G colorimetric measurements resulted in detection limits of 7 and 6 ng/ml, respectively, which were slightly above the MRL of 4 ng/ml. By using a luminescent substrate, however, the MRL could be reached for these antibiotics as well. Based on the results of the individual EIAs rapid multianalyte tests both on microtitre plates and planar microarray chips were developed. Due to altered assay conditions again the systems were optimized and characterized regarding sensitivity and specificity. The sensitivities achieved on the biochip were well comparable with those obtained in the microtitre plate, whereas total assay time could be reduced to 15 min (microtitre plate) and 5 min (biochip), respectively.

Level thesis:degree_level

thesis.doctoral

Grantor dc:publisher

Ludwig-Maximilians-Universität

Year

2003

Author dc:creator

• Strasser, Angelika

Repository record source_url

https://edoc.ub.uni-muenchen.de/1302/

OAI identifier oai:identifier

oai:edoc.ub.uni-muenchen.de:1302