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Competitive immunoassay for analysis of bisphenol A in children’s sera using a specific antibody


Bisphenol A (BPA) has been reported as a potential estrogenic substance that could affect human health and reproduction. In this study, a monoclonal antibody (Mab) against BPA was produced after the immunization of Balb/c mice with a conjugate of 4,4-bis (4-hydroxyphenyl) valeric acid coupling with keyhole limpet hemocyanin (BVA-KLH). The obtained Mab showed higher affinity against BPA and lower cross-reactivity toward 4,4′-sulfonyldiphenol, diphenolic acid, hydroquinone, salicylic acid, and other common phenolic compounds. Basing on the Mab, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) was developed. Under the optimal conditions, the limit of detection (LOD) was found to be 0.1 ng mL−1 with the linear working range of 0.45–10.56 ng mL−1. After sample extraction, the fortified serum samples were detected with intra- and inter-assay recovery ranges of 81.2–92.9 and 84.4–94.4 %, respectively. Then, 100 children’s sera were screened by ic-ELISA. The result showed that 54 % of the serum samples were BPA-positive. The positive samples were purified by immuno-affinity column (IAC) and further confirmed by high-performance liquid chromatography (HPLC) with fluorescence detector measured at λ ex/λ em 228/310 nm in acetonitrile-water solution (v:v, 40:60). The analysis of the unknown samples showed that ic-ELISA agreed well with the HPLC results. It also revealed that the ELISA developed here could be a useful tool for screening BPA in children’s sera before the validation of HPLC.

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This work was financially supported by grants from the National Natural Science Foundation of China (No. 81360429) and Natural Science Foundation of Jiangxi Province (Nos. 20122BAB214006, 20132BAB214004, KJLD14067).

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Correspondence to Renrong Liu.

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The authors declare that they have no competing interests.

Compliance with ethics requirements

All animal studies were carried out in compliance with the administration of affairs approved by the laboratory animal ethics review committee of Jiangxi Province. All the experiments involving human subjects were approved by the committee of Jiangxi Science and Technology Normal University, and informed consent was obtained from all individual participants included in the study.

Additional information

Fanfan Yang, Long Xu and Lixin Zhu contributed equally to this work.

Responsible editor: Philippe Garrigues

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Yang, F., Xu, L., Zhu, L. et al. Competitive immunoassay for analysis of bisphenol A in children’s sera using a specific antibody. Environ Sci Pollut Res 23, 10714–10721 (2016). https://doi.org/10.1007/s11356-016-6231-2

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  • Bisphenol A
  • Monoclonal antibody
  • Sera
  • Immuno-affinity column
  • HPLC