Preparation of polyclonal antibody against PRRSV receptor sialoadhesin

Sialoadhesin(Sn) is a primary receptor of porcine reproductive and respiratory syndrome virus(PRRSV).The specific antibody against Sn is required for investigation of Sn functions during PRRSV infection.To prepare Sn antibody,an epitope gene of Sn,coding 114 amino acids(Sn114) was determined based on the antigenic analysis of complete Sn sequence by DNAstar software.The Sn114 epitope gene sequence was codon-optimized according to E.coli preferred codons and synthesized by a company.The codon-optimized Sn114 gene was cloned into pET-32a(+) plasmid to construct the prokaryotic expression vector of Sn114 gene fused with thioredoxin and His-tag.The expression vector was transformed into E.coli BL21(DE3) to express Sn114 protein with IPTG induction.The Sn114 fusion protein was purified with Ni-NTA agarose beads.The rabbits were immunized with the Sn114 fusion protein,and the antibody titer was measured by ELISA.The result showed that the Sn114 fusion protein was highly expressed in E.coli at,about 63 mg/L.The serum antibody titer reached 1∶10 240 at 34 d after immunization.The macrophage biding assay indicated that the antibody had the ability to specifically detect Sn antigen on porcine alveolar macrophages.