Production of monoclonal antibody against oxytetracycline for developing enzyme-linked immunosorbent assay (ELISA) test kit, 2nd year

Abstract

In Thailand, oxytetracyline (OTC) has been used in the protection and treatment of infected shrimps. The misuse of OTC could lead to OTC residues in shrimps destined for consumption. To prevent consumers from exposure to drug residues and increase of drug resistance pathogens, several food safety authorities in many countries have set the maximum residue limits (MRLs) for OTC and enforced the surveillance detection programs. Therefore, an effective screening method is essential to detect OTC residue in food. Enzyme-linked immunosorbent assay (ELISA) is the suitable methods for screening a large number of samples due to its simplicity, rapidity and cost effectiveness. Consequently, the aim of this study was to develop the methods in order to detect OTC residue in shrimp samples. The results showed that the indirect competitive ELISA using goat anti mouse IgG-hosreradish peroxidase (GAM-HRP) was suitable for OTC detection with the 50% inhibition concentration (IC50) of 5.5 ng/ml and the limit of detection (LOD) of 0.9 ng/ml. The developed ELISA could detect OTC at the concentration between 0.5 ng/ml and 32 ng/ml which covered the current MRLs. In addition, the detection showed strong cross-reactivity to rolitetracycline (RTC) (142%) but did not crossreact with other related antibiotics. When the ELISA was used to detect OTC in fortified shrimp samples, the %recoveries were in the range of 82%-118% for an intra-assay and 96%-113% for an inter-assay. The coefficients of variation of ELISA were 5.5%-13.9% and 5.5%-14.9%, respectively.