Sequencing of the cDNA variable region of a lumpy skin disease virus ORF123 monoclonal antibody and its cross-neutralizing activity analyses against Capripoxvirus members

Abstract

Monoclonal antibody sequencing is crucial for enhancing our understanding of the relationship between antibody neutralization and broad-spectrum binding. Our previous study systematically described a B-cell monoclonal antibody (mAb) derived from lumpy skin disease virus (LSDV) ORF123, which exhibits cross-reactivity with goatpoxvirus (GTPV) and sheeppoxvirus (SPPV). Here, the LSDV ORF123 mAb was sequenced for the first time using the HybSeq HT™ next-generation sequencing technique. Sequencing analyses showed that the variable heavy (V_H) and light (V_L) chains of LSDV ORF123 mAb had significant sequence similarity. Subsequently, the top 1 V_H and V_L of this LSDV ORF123 mAb were synthesized and expressed. Western blotting analyses further identified that the complementary determinant regions (CDR)1, CDR2, and CDR3 are indispensable for antigen-antibody recognition. Crucially, the co-expression of V_H + V_L of this LSDV ORF123 mAb retained its cross-neutralizing activity against LSDV, GTPV, and SPPV. Variable region sequencing of monoclonal antibodies provides a reference for improving the specificity and affinity of potential therapeutic antibody design. The cross-binding and neutralizing activity of the co-expression of V_H + V_L of the LSDV ORF123 mAb strengthens our understanding of the antigenic similarity among different members of the genus Capripoxviruses.