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Preparation of Artificial Antigen and Development of IgY-Based Indirect Competitive ELISA for the Detection of Kanamycin Residues

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Abstract

The present study was aimed to evaluate the feasibility to use chicken egg yolk antibodies (IgY) for the detection of Kanamycin (Kana) residues in food stuffs. Bovine serum albumin (BSA) was conjugated with Kana by carbodiimide method, and the laying chickens were immunized with Kana-BSA conjugate. PEG-6000 method was employed to extract IgY. The peak titer of anti-Kana IgY was 1:256,000 after the fifth booster immunization. The optimal dilution for anti-Kana IgY was 1:25,000 to obtain optical density (OD) of 1.0 at 2 μg/mL of OVA-Kana coating concentration. The indirect competitive ELISA (ic-ELISA) showed that the IC50 value of anti-Kana IgY was 4.48 ng/mL and the regression curve equation was y = −15.62x + 60.17 (R 2 = 0.98, n = 3). The ic-ELISA showed a lower cross-reactivity (less than 0.01 %) with other drugs (except gentamicin—1.91 %). Recoveries from milk, pork, and chicken powder samples were in the range of 82.02 to 98.20 %, with relative standard deviation lower than 5.13 %. Our results indicate that generated anti-Kana IgY can be used in routine screening analysis of Kana residues in food samples.

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Acknowledgments

This work was supported by the Key Construction Program of International Cooperation Base in S&T, Shaanxi Province (2015SD0018), China, Ph.D. Programs Foundation of Ministry of Education of China (20130204110023), Key Construction Program of International Cooperation Base in S&T (2015SD0018), Shaanxi Province, China, and Ministry of Education and State Administration of Foreign Experts Affairs “Overseas Teacher” project (MS2011XBNL057), China.

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Conflict of Interest

Jinxin He declares that he has no conflict of interest. Yuan Wang declares that she has no conflict of interest. Xiaoying Zhang declares that he has no conflict of interest. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted.

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  1. College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, 712100, China

    Jinxin He, Yuan Wang & Xiaoying Zhang

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  1. Jinxin He
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  2. Yuan Wang
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  3. Xiaoying Zhang
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Correspondence to Xiaoying Zhang.

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He, J., Wang, Y. & Zhang, X. Preparation of Artificial Antigen and Development of IgY-Based Indirect Competitive ELISA for the Detection of Kanamycin Residues. Food Anal. Methods 9, 744–751 (2016). https://doi.org/10.1007/s12161-015-0248-x

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  • DOI: https://doi.org/10.1007/s12161-015-0248-x

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