Purification of Porcine Circovirus Type 2 Using an Affinity Chromatography Based on a Neutralizing Monoclonal Antibody against Viral Capsid Protein.

Haiqiao Bian,Chong Yu,Yanwu Wei,Li Feng,Liping Huang,Changming Liu

doi:10.3390/pathogens10121564

Abstract

Porcine circovirus type 2 (PCV2) is a DNA virus without an envelope. The viral particle is icosahedral and has a diameter of approximately 17 nm. In order to obtain the purified virus, a broad-spectrum monoclonal antibody 3A5 against PCV2 was coupled to CNBr-activated SepharoseTM 4B, and an affinity chromatography was established for PCV2 purification. A total of 6.5 mg of purified PCV2a/LG with 97% purity was obtained from 120 mL of the viral culture medium, and only PCV2 was detected by electron microscopy. No significant changes in the antigenic characteristics of the purified virus were detected by a capture enzyme-linked immunosorbent assay (ELISA). Furthermore, the titer of the purified PCV2 was 100 times higher than that of the unpurified virus. This affinity chromatography method was also used to purify PCV2b/LN590516 and PCV2d/SD446F16, and the purified viruses were detected by electron microscopy, capture ELISA, and virus titration, respectively. The results showed that these two strains can be successfully purified, but the yield is lower than that of the PCV2a strain. In addition, the purified virus could be used to study the viral adsorption and invasion of PK15 cells using indirect immunofluorescence assays. A large number of PCV2 signals were detected to transfer from the cellular surface to the periphery of the nucleus of the PK15 cells after 30 min of adsorption of the PCV2 to the PK15 cells. The affinity chromatography is a simple and convenient tool to obtain PCV2 with high purity. It could be applied for virus structure analysis, antibody preparation, and viral adsorption and invasion research.

Highlights

Porcine circovirus type 2 (PCV2) is a member of the genus Circovirus in the familyCircoviridae
The diseases associated with PCV2 infection are defined by the American Porcine Veterinary Association as porcine circovirus-associated disease (PCVAD)
We report an affinity chromatography based on a PCV2 broad-spectrum monoclonal antibody to obtain PCV2 with high purity

Summary

Introduction

Porcine circovirus type 2 (PCV2) is a member of the genus Circovirus in the family. In 1991, the virus was isolated from pigs suffering from post-weaning multisystemic wasting syndrome in Canada [1]. Other diseases are associated with PCV2 infection, including porcine reproductive failure, porcine respiratory disease complex, and granulomatous enteritis. These diseases have caused great economic losses to the pig industry worldwide. The diseases associated with PCV2 infection are defined by the American Porcine Veterinary Association as porcine circovirus-associated disease (PCVAD). In Europe, porcine circovirus disease (PCVD) is often used to refer to PCVAD [2]

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