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Development of an antigen-capture enzyme-linked immunosorbent assay and immunochromatographic strip based on monoclonal antibodies for detection of H6 avian influenza viruses

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Abstract

Continuous surveillance has shown that H6 subtype avian influenza viruses (AIVs) are prevalent in poultry and occasionally break the species barrier to infect humans. It is therefore necessary to establish a specific, rapid and sensitive method to screen H6 AIVs. In this study, a panel of monoclonal antibodies (mAbs) against the hemagglutinin (HA) of an H6 AIV isolate was produced. The purified mAbs have high affinity and specificity for H6 AIVs. An antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) and immunochromatographic strip were developed based on two mAbs (1D7 and 1F12). The AC-ELISA results showed high sensitivity with a limit of detection (LOD) of 3.9 ng/ml for H6 HA protein and 0.5 HAU (HA units)/100 µl for live H6 subtype AIVs. The average recovery of the AC-ELISA with allantoic fluid, respiratory specimens, and cloacal swabs was 91.907 ± 1.559%, 82.977 ± 1.497% and 73.791 ± 2.588%, respectively. The intra- and inter-assay coefficient of variation was less than 10%. The LOD of immunochromatographic strip was 1 HAU when evaluated by the naked eye, and the detection time was less than 10 min without any equipment. Storage at room temperature or 4 °C for 30 days or 60 days had no effect on sensitivity and specificity of the strip. Thus, the AC-ELISA and immunochromatographic strips described here could be a secondary method to diagnose H6 AIV infections with high specificity, sensitivity, and stability.

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Funding

This study was funded by grants from the National Science and Technology Major Project for the Control and Prevention of Major Infectious Diseases in China (2018ZX10711001, 2018ZX10102001 and 2020ZX09001016-004-002), Zhejiang Provincial Natural Science Foundation of China (LY19H260006), and the Independent Task of State Key Laboratory for Diagnosis and Treatment of Infectious Diseases (2019ZZ17).

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Authors and Affiliations

  1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, and National Clinical Research Center for Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University, 79 Qingchun Road, Hangzhou, 310003, Zhejiang, China

    Fan Yang, Yixin Xiao, Fumin Liu, Hangping Yao, Nanping Wu & Haibo Wu

  2. Animal Husbandry and Veterinary Institute, Zhejiang Academy of Agricultural Science, Hangzhou, 310021, China

    Lihua Xu

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  1. Fan Yang
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  2. Yixin Xiao
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Correspondence to Haibo Wu.

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The animal experiment was approved by the First Affiliated Hospital, School of Medicine, Zhejiang University (No. 2017-015).

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Yang, F., Xiao, Y., Xu, L. et al. Development of an antigen-capture enzyme-linked immunosorbent assay and immunochromatographic strip based on monoclonal antibodies for detection of H6 avian influenza viruses. Arch Virol 165, 1129–1139 (2020). https://doi.org/10.1007/s00705-020-04602-w

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  • DOI: https://doi.org/10.1007/s00705-020-04602-w

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