Optimization of prokaryotic expression condition of SARS-CoV-2 papain-like protease and preparation and identification of polyclonal antibody

  • LIU Zhicheng ,
  • YAN Gang’an ,
  • YAN Haohao ,
  • LIU Xiaoli ,
  • LIU Xiaoping ,
  • CHEN Yunyu
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  • Institute for Drug Screening and Evaluation, Wannan Medical College, Wuhu 241000, China

Received date: 2023-04-13

  Online published: 2024-01-09

Abstract

Objective: To optimize the expression conditions of SARS-CoV-2 papain-like protease (papain-like protease, PLpro) in Escherichia coli (E.coli) and prepare highly specific polyclonal antibody against rat PLpro. Methods: The optimal expression conditions of PLpro in E.coli were determined by optimizing induction time, induction temperature and IPTG concentration. After PLpro being isolated and purified by HisTrapTM affinity chromatography column, the rat was immunized with PLpro as antigen to prepare anti-PLpro polyclonal antibody. The titer, antigen specificity and sensitivity of polyclonal antibody were determined by enzyme-linked immunosorbent assay (ELISA) and Western Blot experiments. Results: The optimum induction temperature of PLpro was 20 ℃, the induction time was 10 h, and the concentration of IPTG was 0.2 mmol/L. The titer of anti-PLpro polyclonal antibody reached to 1∶256 000 with good antigen specificity, and the detection limit of anti-PLpro polyclonal antibody was 12.5 ng with good sensitivity. Conclusion: The expression conditions of PLpro in E.coli was successfully optimized and highly specific polyclonal antibody against rat PLpro successfully prepared, which laid a foundation for the study of the immunological function of PLpro in COVID-19.

Cite this article

LIU Zhicheng , YAN Gang’an , YAN Haohao , LIU Xiaoli , LIU Xiaoping , CHEN Yunyu . Optimization of prokaryotic expression condition of SARS-CoV-2 papain-like protease and preparation and identification of polyclonal antibody[J]. Journal of Baotou Medical College, 2024 , 40(1) : 85 -90 . DOI: 10.16833/j.cnki.jbmc.2024.01.015

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