Preparation and identification of polyclonal antibodies specific for nsp4 protein of porcine reproductive and respiratory syndrome virus.

Author: Xinna CAI ¹ ; Min TAN ¹ ; Shengliang CAO ¹ ; Yan HUANG ¹ ; Fachao SUN ¹ ; Yingli SHANG ¹ ; Sidang LIU ¹ ; Yihong XIAO ¹
Author Information

1. College of Veterinary Medicine and Animal Sciences, Shandong Agricultural University, Tai'an 271000, Shandong, China.
Publication Type:Journal Article
Keywords: identification; nsp4; polyclonal antibody; porcine reproductive and respiratory syndrome virus
From: Chinese Journal of Biotechnology 2017;33(8):1276-1283
CountryChina
Language:Chinese
Abstract: To obtain specific antibodies against nsp4 protein of porcine reproductive and respiratory syndrome virus (PRRSV), nsp4 gene was amplified by RT-PCR and cloned into pET-28a(+) vector, designated pET28a-nsp4. pET28a-nsp4 was transformed into Escherichia coli Trasseta (DE3) cells and expressed after induction of IPTG. SDS-PAGE analysis showed that the recombinant protein was expressed in soluble form with the molecular weight of 26 kDa. The soluble fusion protein in the supernatant was purified using Ni+-NTA affinity chromatography. New Zealand rabbits were immunized by the purified nsp4 and anti-sera against nsp4 were obtained. The titer of polyclonal antibodies was about 106 and showed good specificity and sensitivity in the immunofluorescence assay and Western blotting analysis. The polyclonal antibodies also recognized native nsp4 form PRRSV infected Marc-145 cells, providing a useful tool in PRRSV replication mechanism study.