Journal
FOOD AND AGRICULTURAL IMMUNOLOGY
Volume 27, Issue 3, Pages 314-328Publisher
TAYLOR & FRANCIS LTD
DOI: 10.1080/09540105.2015.1089843
Keywords
tylosin; tilmicosin; monoclonal antibody; ELISA; immunochromatographic strip
Funding
- Key Programs from MOST [2012BAD29B05, 2012BAD29B04]
- Natural Science Foundation of Jiangsu Province
- MOF
- MOE [BK20140003, 201310135]
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Tylosin (TYL) and tilmicosin (TIM) are macrolide antibiotics, and maximum residue limits (MRLs) have been set to control their illegal usage in food. We developed a sensitive indirect competitive enzyme-linked immunosorbent assay and rapid immunochromatographic strip (ICS) test to simultaneously detect TYL and TIM based on an innovative hapten (TYL-carboxymethoxylamine hemihydrochloride). The monoclonal antibody 2B3 was obtained with the isotype IgG1, and TYL and TIM had half maximal inhibitory concentrations of 0.57 and 2.10ng/ml, respectively. The cross-reactivity values were 100%, 27.57%, 97.43%, and 62.42% for TYL, TIM, desmycosin, and acetylisovaleryl tylosin, respectively. By optimizing, the standard dilution buffer was based on 0.8% NaCl, pH 7.4 and 5% acetonitrile. The recoveries ranged from 89.37% to 112.00% in the honey samples, which suggests that the assay would be reliable in the analysis of real honey samples. In addition, an ICS was developed for qualitative, semi-quantitative, and spot detection analysis, with cut-off values of 5 and 15ng/ml for TYL and TIM in phosphate-buffered saline, and 10 and 20ng/ml for TYL and TIM in honey, respectively. The results show that these assays can be used to analyze real samples with the strictest MRL being 50ng/ml for both TYL and TIM.
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