Production and application of mouse monoclonal antibodies targeting linear epitopes in pB602L of African swine fever virus

Pengfei Wang,Lili Wei,Zhibin Shi,Shida Wang,Chunguo Liu,Zaisi Liu,Ming Wang,Decheng Yang,Jingfei Wang,Zhenzhao Sun,Lili Wen,Xijun He,Yue Chi

doi:10.1007/s00705-021-05335-0

Abstract

African swine fever (ASF) is an acute hemorrhagic disease of domestic pigs. The causative agent of ASF, ASF virus (ASFV), is a double-stranded DNA virus, the sole member in the family Asfarviridae. The non-structural protein pB602L of ASFV is a molecular chaperone of the major capsid protein p72 and plays a key role in icosahedral capsid assembly. This protein is antigenic and is a target for developing diagnostic tools for ASF. To generate monoclonal antibodies (mAbs) against pB602L, a prokaryotically expressed recombinant pB602L protein was produced, purified, and used as an antigen to immunize mice. A total of eight mouse mAbs were obtained, and their binding epitopes were screened by Western blot using an overlapping set of polypeptides from pB602L. Three linear epitopes were identified and designated epitope 1 (366ANRERYNY373), epitope 2 (415GPDAPGLSI423), and epitope 3 (498EMLNVPDD505). Based on the epitope recognized, the eight mAbs were placed into three groups: group 1 (B2A1, B2F1, and B2D10), group 2 (B2H10, B2B2, B2D8, and B2A3), and group 3 (B2E12). The mAbs B2A1, B2H10, and B2E12, each representing one of the groups, were used to detect pB602L in ASFV-infected porcine alveolar macrophages (PAMs) and pig tissues, using an indirect fluorescence assay (IFA) and immunohistochemical staining, respectively. The results showed that pB602L was detectable with all three mAbs in immunohistochemical staining, but only B2H10 was suitable for detecting the proteins in ASFV-infected PAMs by IFA. In summary, we developed eight anti-pB602L mouse mAbs recognizing three linear epitopes in the protein, which can be used as reagents for basic and applied research on ASFV.

Highlights

African swine fever (ASF) is an acute hemorrhagic disease of domestic pigs
The expression and purity of the recombinant pB602L protein were first assessed by SDS-PAGE
The expressed pB602L was identified by Western blot assays using an anti-GST monoclonal antibody (Fig. 2B) and an anti-ASF virus (ASFV) pig serum (Fig. 2C)

Summary

Introduction

African swine fever (ASF) is an acute hemorrhagic disease of domestic pigs. This disease often results in devastating economic losses to the pig industry of affected countries because of its high rate of morbidity and mortality, and it is listed as a notifiable disease by the World Organization for Animal Health (OIE). In 2018, the first ASF outbreak was reported in Liaoning Province in China [11, 28]. The disease has occurred in almost every major pig-production area of China [25]. Recent surveillance has identified naturally attenuated ASFV strains in several provinces of China, making the control situation even more complex [23]. The disease has been reported recently in other Asian countries [17]

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Conclusion