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Development of Competitive ELISA and CLEIA for Quantitative Analysis of Polymyxin B

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Abstract

Polymyxin B (PMB), a member of polypeptide antibiotics, is widely used for the treatment of infection in animals such as cattle, sheep, pigs, and chickens. However, it is toxic on the kidneys and nervous system, and polymyxin resistance is increasingly reported, which leaves a serious threat to human health. Therefore, it is essential to establish rapid methods for detecting PMB with high sensitivity and specificity. In this study, an anti-PMB polyclonal antibody (pAb) was obtained by immunizing New Zealand white rabbits with PMB conjugated with glycosylated bovine serum albumin (GBSA). Indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and indirect competitive chemiluminescent enzyme immunoassay (ic-CLEIA) were developed. Under the optimal conditions, inhibitory concentrations (IC50) of PMB were 257.1 ng/mL (ic-ELISA) and 250.8 ng/mL (ic-CLEIA); the limits of detection (LOD) were 17.4 ng/mL (ic-ELISA) and 14.5 ng/mL (ic-CLEIA), respectively. Cross-reactivity of the pAb toward polymyxin E (PME) was 257.1%, and no response was found with other antibiotics. The recovery rates in spiked meat samples were 77.4~106.1% (ic-ELISA) and 84.1~107.1% (ic-CLEIA), respectively.

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Funding

This work was supported by the Key Program for International S&T Cooperation Project of Shaanxi Province (2017KW-ZD-10), the strategic program UID/BIA/04050/2013 (POCI-01-0145-FEDER-007569) funded by national funds through the FCT I. P., the Ministério da Ciência, Tecnologia e Ensino Superior (MCTES), the ERDF through the COMPETE2020 - Programa Operacional Competitividade e Internacionalização (POCI), and the Ministry of Education and Science of the Russian Federation, unique project identifier RFMEFI60417X0198.

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Authors and Affiliations

  1. Centre of Molecular and Environmental Biology, Department of Biology, University of Minho, Campus de Gualtar, 4710-057, Braga, Portugal

    Long Xu, Alberto C. P. Dias & Xiaoying Zhang

  2. Department of Immunology, I. Mechnikov Research Institute of Vaccines and Sera, Moscow, Russia, 105064

    Maksim Burkin

  3. Faculty of Chemistry, M.V. Lomonosov Moscow State University, Moscow, Russia, 119991

    Sergei Eremin

  4. College of Veterinary Medicine, Northwest Agriculture and Forestry University, PostBox 19, Xinong Rd. 22, Yangling, 712100, Shaanxi Province, China

    Xiaoying Zhang

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  1. Long Xu
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  2. Maksim Burkin
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  4. Alberto C. P. Dias
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Correspondence to Xiaoying Zhang.

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Conflict of Interest

Long Xu declares that he has no conflict of interest. Maksim Burkin declares that he has no conflict of interest. Sergei Eremin declares that he has no conflict of interest. Alberto C. P. Dias declares that he has no conflict of interest. Xiaoying Zhang declares that he has no conflict of interest.

Ethical Approval

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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Informed consent was obtained from all individual participants included in the study (in case humans are involved).

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Xu, L., Burkin, M., Eremin, S. et al. Development of Competitive ELISA and CLEIA for Quantitative Analysis of Polymyxin B. Food Anal. Methods 12, 1412–1419 (2019). https://doi.org/10.1007/s12161-019-01477-9

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  • DOI: https://doi.org/10.1007/s12161-019-01477-9

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