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Selective Immunodetection of Difloxacin in Animal Muscles and Sera: Role of Hapten Orientation

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Abstract

The influence of sarafloxacin (SAR) hapten orientation in immunogen on antibody specificity was examined. The spatial orientation of SAR linked through its carboxyl group to the carrier resulted to SAR-selective response (Anal Methods 2016, 8:5843–5850). To provide opposite orientation of SAR in immunogen, it was linked by its secondary amine to (1) succinic anhydride-modified BSA using carbodiimide-mediated conjugation and to (2) unmodified BSA using formaldehyde condensation method. Two versions of indirect competitive enzyme-linked immunosorbent assays (ELISA) based on generated antibodies were developed. A panel of homo- and heterologous conjugates was examined as potential coating antigens. No cross-reactions were registered in ELISA-1 and ELISA-2 except for fluorophenyl-containing fluoroquinolones difloxacin (DIF), SAR, and tosufloxacin as 138, 100 and 25% and 175, 100 and 8%, respectively. Among the recognized analytes, only DIF is authorized in meat products, so the developed tests were selective for DIF determination in bovine and porcine muscles and sera samples. The values of half-maximal inhibition concentration (IC50) for these assay versions were estimated to be 1.2 and 0.25 ng/ml. DIF detection limits (IC10) were, respectively, 0.15 and 0.015 ng/ml that allowed measuring of MRL and ten times lower level of DIF in the animal muscles. The developed ELISAs were also suitable for DIF determination in animal sera in the concentration range 1000–1 ng/ml.

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Authors and Affiliations

  1. I. Mechnikov Research Institute of Vaccines and Sera, Moscow, 105064, Russia

    Inna A. Galvidis & Maksim A. Burkin

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  1. Inna A. Galvidis
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  2. Maksim A. Burkin
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Correspondence to Maksim A. Burkin.

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Inna Galvidis declares that she has no conflict of interest. Maksim Burkin declares that he has no conflict of interest.

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This article does not contain any studies with human participants performed by any of the authors. All studies with experimental animals were carried out under the guidelines of Ethics Committee of I. Mechnikov Research Institute of Vaccines and Sera.

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Galvidis, I.A., Burkin, M.A. Selective Immunodetection of Difloxacin in Animal Muscles and Sera: Role of Hapten Orientation. Food Anal. Methods 10, 1755–1764 (2017). https://doi.org/10.1007/s12161-016-0730-0

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  • DOI: https://doi.org/10.1007/s12161-016-0730-0

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