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Magnetic-assisted biotinylated single-chain variable fragment antibody-based immunoassay for amantadine detection in chicken

Analytical and Bioanalytical Chemistry volume 410, pages 6197–6205 (2018)Cite this article

Abstract

A sensitive competitive immunoassay with simple operation was developed for the detection of the anti-virus drug amantadine (AMD). The single-chain variable fragment (scFv) antibody against AMD was site-specific biotinylated and overexpressed as a secreted body in Escherichia coli AVB101. Horseradish peroxidase-labeled streptavidin-biotinylated scFv antibody (HRP-SA-BIO-scFv) could specifically bind to AMD-functionalized magnetic beads (MBs) and then the immune complexes were separated from the matrix solution by magnet. The concentration of the AMD could be known by the measurement of the signal produced by the horseradish peroxidase. The newly established assay provides a significant improvement in comparison to the conventional ELISA without SA-BIO signal amplification and MBs separation. The limit of detection and assay time was 0.64 vs. 8.4 ng/mL and 50 vs. 150 min, respectively. The recoveries ranged from 77.8 to 112% with the coefficient of variation less than 13%. The immunoassay exhibited an obvious cross-reactivity to rimantadine (84%), 1-(1-adamantyl)ethylamine (72%), and somantadine (63%). These results demonstrated that the developed immunoassay provided a sensitive, rapid, and accurate approach for the detection of AMD in chicken by employing MBs as solid phase and SA-BIO as signal amplification. When applied in natural chicken samples, the newly established method provided results consistent with those from UPLC-MS/MS, suggesting that the proposed method could be used for rapid screening of the target of interest; the new immunoassay could also be extended to other small molecular contaminants and thus represents a universal strategy for food safety analysis.

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Acknowledgments

This work was supported by grants from the National Natural Science Foundation of China (No. 31472236 and 31672600).

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Authors and Affiliations

  1. Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Veterinary Medicine, China Agricultural University, Beijing, 100193, People’s Republic of China

    Sanlei Xie, Kai Wen, Jie Xie, Tao Peng, Jianyi Wang, Kai Yao, Shuangyang Ding & Haiyang Jiang

  2. Beijing Key Laboratory of Detection Technology for Animal-Derived Food Safety, Beijing Laboratory for Food Quality and Safety, Beijing, 100193, People’s Republic of China

    Sanlei Xie, Kai Wen, Jie Xie, Tao Peng, Jianyi Wang, Kai Yao, Shuangyang Ding & Haiyang Jiang

  3. Department of Mechanical and Electrical Engineering, College of Engineering, China Agricultural University, Beijing, 100083, China

    Yongjun Zheng

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  1. Sanlei Xie
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Correspondence to Haiyang Jiang.

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Xie, S., Wen, K., Xie, J. et al. Magnetic-assisted biotinylated single-chain variable fragment antibody-based immunoassay for amantadine detection in chicken. Anal Bioanal Chem 410, 6197–6205 (2018). https://doi.org/10.1007/s00216-018-1227-z

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  • DOI: https://doi.org/10.1007/s00216-018-1227-z

Keywords

  • Immunoassay
  • Biotinylated scFv
  • Magnetic beads
  • Amantadine
  • Chicken