Development of whole-porcine monoclonal antibodies with potent neutralization activity against classical swine fever virus (CSFV) from single B cells

Abstract

Classical swine fever (CSF) is a highly contagious swine disease found worldwide that has caused devastating economic losses. However, there are few efficacious mAbs against the CSF virus (CSFV) that can be used for treatment because most mAbs against CSFV are derived from mouse hybridoma cells and these murine mAbs have disadvantages of inefficient effector functions elicitations and high immunogenicity in vivo. Accordingly, we characterized whole-porcine anti-CSFV neutralizing mAbs (NAbs) isolated directly from single B cells sorted from a CSFV-vaccinated pig using the fluoresceinated conserved linear neutralizing epitope of the CSFV E2 protein and fluorophore conjugated goat anti-pig IgG. Immunoglobulin (Ig) genes were isolated via nested PCR, and two porcine mAbs termed HK24 and HK44 were produced. We determined that these mAbs can bind to E2 protein and recognize sites within this major antigenic epitope. In addition, we found that mAbs HK24 and HK44 exhibit potent neutralizing activity against CSFV, and they can protect PK-15 cells from infections in vitro with potent IC50 values of 9.3 g/ml and 0.62 g/ml, respectively. Notably, we demonstrated that these two mAbs can be used as novel reagents for detecting virus infection. These data suggest that our results not only provide a method for efficiently obtaining mAbs against CSFV but also offer promising mAb candidates for development of antibody-based diagnostic and antiviral agents.\n\nImportanceNeutralizing monoclonal antibodies (NAbs) can prevent and may slow the spread of virus infection. The discovery of NAbs that recognize classical swine fever virus (CSFV) necessitates new technologies because the NAbs produced by immunization and hybridoma technology could not be transferred to in vivo research. Multiple full-length human therapeutic antibodies have been produced via single-cell polymerase chain reactions but whole-porcine NAbs for CSFV have not been generated. In this study, two whole-porcine mAbs, named HK24 and HK44, were isolated from epitope-specific single B cells. We demonstrate that these two mAbs have potent neutralizing activity against CSFV and can protect cells against viral infection. Therefore, they may facilitate the development of vaccines or antiviral drugs that offer the advantages of stability and low immunogenicity.