Anti-AOZ monoclonal antibody, clone W74K74

The use of furazolidone in food animals has been banned in European Union (EU) because of its carcino-genicity and mutagenicity on human health, but its continued misuse is widespread. Therefore, there is an urgent need for a simple, reliable, and rapid method for the detection of its marker residue, 3-amino-2-oxazolidinone (AOZ), in food products. In this regard, a sensitive and reliable electrochemical method was presented to detect AOZ based on a novel label-free electrochemical impedimetric immunosensor to address this need. The immobilization of monoclonal antibody against AOZ (denoted as AOZ-McAb) on the gold electrode was carried out through a stable acyl amino ester intermediate generated by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydrosuccinimide (NHS), which could condense antibodies on the self-assembled monolayer (SAM). The detection of AOZ was performed by measuring the relative change in charge transfer resistance before and after AOZ and AOZ-McAb immunoreaction by electrochemical impedance spectroscopy (EIS). Under the optimized conditions, the relative change in charge transfer resistance was proportional to the logarithmic value of AOZ concentrations in the range of 20.0 to 1.0 x 10(4) ng mL(-1) (r = 0.9987). Moreover, the proposed immunosensor has a high selectivity to AOZ alone with no significant response to the metabolites of other nitrofuran antibiotics, such as 3-amino-5-morpholinomethyl-2-oxazolidinone (AMOZ), semicarbazide (SEM), and 1-aminohydantoin hydrochloride (AHD). This protocol has been applied to detect AOZ in food samples with satisfactory results. Crown Copyright (C) 2011 Published by Elsevier B.V. All rights reserved.

We presented the validation method of the quantitative determination of Nitrofuran (AOZ) and Chloramphenicol in honey samples by the RidaScreen (R) test system (according to Commission Decision 2002/657/EC and Guidelines for the validation of screening methods for residues of veterinary medicines (initial validation and transfer) of 20/01/2010). We revealed several validation indicators like cc ss (detection capability), accuracy (precision), recovery and some others during the initial validation (specificity and selectivity). We determined the minimum detection values of Nitrofuran (AOZ) in honey, the highest value (the highest response) for blank samples is 0.066 mu g/kg and the lowest value for enriched samples (lowest response) is 0.401 mu g/kg. Based on the obtained results none of the responses for enriched samples do not coincide with the range of replies of blank samples. Hence, it follows that the detection capability (CC ss) for this screening method is less than or equal to 0.4 mu g/kg. For the determination of Chloramphenicol, the highest response is noted for blank samples - 0.021 mu g/kg (21 ng/kg) and the lowest response noted for enriched samples is 0.048 mu g/kg (48 ng/kg). None of the responses for enriched samples do not coincide with the range of replies of blank matrix (samples), therefore we could conclude that the CC ss of this screening method is less than or equal to 0.05 mu g/kg (ss-error < 5 %) and the cut-off value of this test is 0.048. According to the validation the lowest value of substances that could detected by ELISA using the following test systems for competitive ELISA (enzyme-linked immunoassay) for Nitrofuran (AOZ) - 0.4 mu g/kg, for Chloramphenicol - 0.05 mu g/kg. Described in State Standard 4497: 2005 "Natural Honey. Technical conditions" the maximum allowable level for Chloramphenicol - 0.3 mu g/kg and Nitrofuran (AOZ) - 0.6 mu g/kg require correction according to the established validation parameters.