非洲猪瘟病毒pE120R蛋白单克隆抗体的制备

doi:10.11843/j.issn.0366-6964.2024.01.036

Abstract

Abstract: This study aimed to express the African swine fever virus (ASFV) capsid protein pE120R and prepare the monoclonal antibody (mAb) for the further study of its function. The pE120R gene was amplified from pCAGGS-Flag-pE120R and the prokaryotic expression plasmid pGEX-6p1-pE120R was constructed. After transforming into BL21(DE3)competent cells, it was induced to express soluble pE120R protein by IPTG. pE120R protein was purified by GST Beads and immunized BALB/c mice, the spleen cells of the immunized mice were collected and perform cell fusion with SP2/0 cell, thus a hybridoma cell line secreting pE120R monoclonal antibody was obtained by indirect ELISA. The results showed that the prokaryotic expression plasmid pGEX-6p1-pE120R was successfully constructed and the high purity pE120R protein was obtained. The results of Western blot and indirect immunofluorescence assay (IFA) showed that pE120R mAb could specifically recognize Flag-pE120R protein expressed by HEK293T cells transfected with plasmid and pE120R protein expressed by ASFV infected alveolar macrophage cells (PAMs). The binding site of this mAb is in the 30-60 aa region, and the mAb subclass identifies the heavy chain as IgG2a. In this study, the soluble pE120R protein was successfully expressed and purified; the monoantibody against pE120R was prepared, which has good specificity and laid a foundation for exploring the biological function of ASFV pE120R protein.

Key words: African swine fever virus, pE120R protein, prokaryotic expression, monoclonal antibody

CLC Number:

S852.651

LIU Chuanxia, WANG Xiao, LI Xuewen, BAO Miaofei, LI Tingting, CHEN Xin, WENG Changjiang, ZHENG Jun. Preparation of Monoclonal Antibody of African Swine Fever Virus pE120R[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 388-394.

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Preparation of Monoclonal Antibody of African Swine Fever Virus pE120R

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