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Electrochemical mixed aptamer-antibody sandwich assay for mucin protein 16 detection through hybridization chain reaction amplification

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Abstract

A mixed aptamer-antibody sandwich assay for the determination of mucin protein 16 (MUC16) was developed based on hybridization chain reaction (HCR) with methylene blue (MB) as an electrochemical indicator. First, MUC16 antibody was adsorbed onto the surface of the Au nanoparticle (AuNP)-modified indium tin oxide (ITO) electrode to effectively capture the target MUC16. After MUC16 was captured by the MUC16 aptamer, an antibody/MUC16/aptamer sandwich structure formed for the highly selective detection of MUC16. The 3′ end of the aptamer was then subjected to HCR with the assistance of auxiliary probes to obtain DNA concatemers. Numerous MB molecules bonded with G bases in the DNA concatemers by immersing the modified ITO electrode into a stirred solution containing MB with KCl. Stepwise changes in the microscopic features of the electrode surface were studied by scanning electron microscopy (SEM). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the electrochemical behavior of the different modified electrodes. The oxidation current of MB was detected by differential pulse voltammetry (DPV). Under the optimum conditions, the proposed mixed aptamer-antibody sandwich assay showed wide dynamic range from 0.39 to 200 unit mL−1 with a low detection limit of 0.02 unit mL−1 (S/N ratio = 3). The proposed method showed good accuracy, selectivity, and acceptable reproducibility.

Graphical abstract

An electrochemical mixed aptamer-antibody sandwich assay based on the aptamer-induced HCR amplification strategy was fabricated for the highly sensitive detection of MUC16. The mixed aptamer-antibody sandwich assay showed acceptable performance of detection range, detection limit, reproducibility, and selectivity.

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Funding

This research was financially supported by the Medical Health Science and Technology Project of Zhejiang Provincial Health Commission (No. 2017PY024 and No. 2018PY022), the Zhejiang Provincial Natural Science Foundation of China (LQ19H090019), and the Public Welfare from the Science and Technology Department of Zhejiang Province (No. 2017C33201).

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Author notes

  1. Lingsong Lu and Bei Liu contributed equally to this work.

Authors and Affiliations

  1. Department of Central Laboratory, Affiliated Hangzhou First People’s Hospital, Zhejiang University School of Medicine, Hangzhou, 310006, Zhejiang, China

    Lingsong Lu, Jianhang Leng, **ao Ma & Huihui Peng

  2. Department of Reproductive Genetics, Women’s Hospital, Zhejiang University School of Medicine, Hangzhou, 310006, Zhejiang, China

    Bei Liu

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  1. Lingsong Lu
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  2. Bei Liu
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Correspondence to Lingsong Lu.

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All procedures performed in this study involving human participants were in accordance with the ethical standards of the 1964 Declaration of Helsinki and its later amendments or comparable ethical standards, and were approved by the Ethical Committee of Hangzhou First People’s Hospital. Informed consent was obtained from all individual participants included in the study.

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Lu, L., Liu, B., Leng, J. et al. Electrochemical mixed aptamer-antibody sandwich assay for mucin protein 16 detection through hybridization chain reaction amplification. Anal Bioanal Chem 412, 7169–7178 (2020). https://doi.org/10.1007/s00216-020-02849-5

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  • DOI: https://doi.org/10.1007/s00216-020-02849-5

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