Development of a sandwich ELISA for detecting 3AB non-structural protein of foot-and-mouth disease virus.

Yuanfang FU; Wei HE; Pu Sun; Lin Yang; Huifang Bao; Yimei Cao; Xingwen Bai; Pinghua LI; Dong LI; Yingli Chen; Lei Liu; Zengjun LU; Zaixin Liu

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Development of a sandwich ELISA for detecting 3AB non-structural protein of foot-and-mouth disease virus.

Author: Yuanfang FU ¹ ; Wei HE ¹ ; Pu SUN ¹ ; Lin YANG ¹ ; Huifang BAO ¹ ; Yimei CAO ¹ ; Xingwen BAI ¹ ; Pinghua LI ¹ ; Dong LI ¹ ; Yingli CHEN ¹ ; Lei LIU ² ; Zengjun LU ¹ ; Zaixin LIU ¹
Author Information

1. State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Lanzhou 730046, Gansu, China.
2. College of Veterinary Sciences, Gansu Agricultural University, Lanzhou 730070, Gansu, China.
Publication Type:Journal Article
Keywords: foot-and-mouth disease; non-structural proteins; quantitative ELISA; vaccine antigen purity
MeSH: Animals; Antibodies, Viral; Enzyme-Linked Immunosorbent Assay; Foot-and-Mouth Disease/prevention & control*; Foot-and-Mouth Disease Virus; Viral Nonstructural Proteins/genetics*; Viral Vaccines
From: Chinese Journal of Biotechnology 2020;36(11):2357-2366
CountryChina
Language:Chinese
Abstract: Antigenic purity is important for quality control of the foot-and-mouth (FMD) whole virus inactivated vaccine. The recommended method for evaluation the antigenic purity of FMD vaccine is to check the serum conversion to non-structural protein (NSP) 3AB antibody after 2 to 3 times inoculation of animals with inactivated vaccine. In this study, we developed a quantitative ELISA to detect the amount of residual 3AB in vaccine antigen, to provide a reference to evaluate the antigenic purity of FMD vaccine. Monoclonal antibody (Mab) of NSP 3A and HRP-conjugated Mab of NSP 3B were used to establish a sandwich ELISA to quantify the NSP 3AB in vaccine antigen of FMD. Purified NSP 3AB expressed in Escherichia coli was serially diluted and detected to draw the standard curve. The detectable limit was determined to be the lowest concentration of standard where the ratio of its OD value to OD blank well was not less than 2.0. Results: The OD value was linearly corelated with the concentration of 3AB protein within the range between 4.7 and 600 ng/mL. The correlation coefficient R² is greater than 0.99, and the lowest detectable limit is 4.7 ng/mL. The amount of 3AB protein in non-purified inactivated virus antigen was detected between 9.3 and 200 ng/mL depending on the 12 different virus strains, whereas the amount of 3AB in purified virus antigen was below the lowest detectable limit. The amount of 3AB in 9 batches of commercial FMD vaccine antigens was between 9.0 and 74 ng/mL, whereas it was below the detectable limit in other 24 batches of commercial vaccine antigens. Conclusion: the sandwich ELISA established in this study is specific and sensitive to detect the content of 3AB protein in vaccine antigen of FMD, which will be a useful method for evaluation of the antigenic purity and quality control of FMD inactivated vaccine.