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

Pathogens ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 1564
Author(s):  
Haiqiao Bian ◽  
Chong Yu ◽  
Yanwu Wei ◽  
Li Feng ◽  
Changming Liu ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibody ◽  
Affinity Chromatography ◽  
Porcine Circovirus Type ◽  
Porcine Circovirus Type 2 ◽  
Porcine Circovirus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Chromatography Method ◽  
Viral Adsorption

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.

scholarly journals Development and Application of a Competitive Enzyme-Linked Immunosorbent Assay for the Detection of Serum Antibodies to Porcine Circovirus Type 2

2000 ◽  
Vol 12 (5) ◽  
pp. 400-405 ◽  
Author(s):  
Ian W. Walker ◽  
Carrie A. Konoby ◽  
Victoria A. Jewhurst ◽  
Irene McNair ◽  
Francis McNeilly ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Large Scale ◽  
Porcine Circovirus Type ◽  
Postweaning Multisystemic Wasting Syndrome ◽  
Porcine Circovirus Type 2 ◽  
Porcine Circovirus ◽  
Enzyme Linked Immunosorbent Assay ◽  
The Mean ◽  
Immunofluorescent Test

We report the development of a competitive enzyme-linked immunosorbent assay (c-ELISA) for the detection of antibodies to porcine circovirus type 2 (PCV2), the agent associated with the recently described postweaning multisystemic wasting syndrome in pigs. At present, no method has been published describing a c-ELISA for the detection of antibodies to PCV2, and currently employed tests are impractical for use in some laboratories. The assay described here uses a cell culture isolate of porcine circovirus type 2 as antigen and a PCV2-specific monoclonal antibody as the competing reagent. Evaluation of the ELISA was performed by comparison with results obtained using an indirect immunofluorescent test on 484 sera from pig herds in the United Kingdom, Canada, France, and the USA and serial bleeds from pigs experimentally infected with porcine circoviruses. The sensitivity and specificity of the ELISA were determined as 99.58% and 97.14%, respectively, at 2 standard deviations (SD) from the mean or 95.81% and 100% at 3 SD from the mean. Using this ELISA, a serologic survey of 461 sera collected from commercial pig herds in Northern Ireland between 1973 and 1999 was undertaken. Analysis of the results of this survey demonstrated that the number of ELISA-positive sera detected in an individual year during this period ranged from 55% to 100%. This c-ELISA has applications for large-scale rapid diagnosis of PCV2 infection in pig populations worldwide and for immunoscreening of sera from other species for antibodies to PCV2.

scholarly journals Effects of Porcine Circovirus Type 2 (PCV2) Maternal Antibodies on Experimental Infection of Piglets with PCV2

2005 ◽  
Vol 12 (11) ◽  
pp. 1347-1351 ◽  
Author(s):  
N. E. McKeown ◽  
T. Opriessnig ◽  
P. Thomas ◽  
D. K. Guenette ◽  
F. Elvinger ◽  
...  
Keyword(s):  
Porcine Circovirus Type ◽  
Porcine Circovirus Type 2 ◽  
Pcv2 Infection ◽  
Maternal Antibodies ◽  
Porcine Circovirus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Group A ◽  
Group B ◽  
Group D

ABSTRACT To determine the effects of porcine circovirus type 2 (PCV2) maternal antibodies on and response to experimental PCV2 infection, 24 piglets were divided into four groups on the basis of the enzyme-linked immunosorbent assay titers of PCV2 maternal antibodies: group A (n = 6; sample/positive [S/P] ratio, <0.2), group B (n = 5; S/P ratio, >0.2 to <0.5), and groups C (n = 8) and D (n = 5) (S/P ratio, >0.5). Piglets in groups A, B, and C were inoculated with PCV2 at day 0 and challenged with PCV2 at day 42. Group D piglets were not exposed to PCV2 at day 0 but were challenged at day 42. Before challenge, seroconversion to PCV2 antibodies occurred in five of six group A piglets, and the antibody level rose above the cutoff level in one of five group B piglets. Viremia was detected in five of six, four of five, and two of eight pigs in groups A, B, and C, respectively. After challenge, PCV2 DNA was detectable from 7 to 21 days postchallenge in the sera from six of six, four of five, three of eight, and five of five pigs in groups A, B, C, and D, respectively. The results indicated that protection against PCV2 infection conferred by maternal antibodies is titer dependent: higher titers are generally protective, but low titers are not.

scholarly journals Cross-Sectional Study on the Sero- and Viral Dynamics of Porcine Circovirus Type 2 in the Field

Vaccines ◽  
2020 ◽  
Vol 8 (2) ◽  
pp. 339
Author(s):  
Chao-Nan Lin ◽  
Ni-Jyun Ke ◽  
Ming-Tang Chiou
Keyword(s):  
Porcine Circovirus Type ◽  
Porcine Circovirus Type 2 ◽  
Porcine Circovirus ◽  
Current Status ◽  
Economic Losses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Viral Dynamics ◽  
Limited Information ◽  
Window Period

Porcine circovirus-associated diseases (PCVADs) cause considerable economic losses in industrial pork production in the field. To minimize the economic losses due to PCVAD, porcine circovirus type 2 (PCV2) vaccines have been developed, and there is widespread vaccination worldwide today. However, limited information is available concerning the current status of PCV2 infection in the field on the Asian continent. The present study aimed to assess sero- and viral dynamics of PCV2 from 12 PCV2-contaminated pig herds with vaccination against PCV2 in Southern and Central Taiwan. In particular, the level of PCV2 load during the window period for seroconversion using real-time polymerase chain reaction and a commercial enzyme-linked immunosorbent assay (ELISA) kit. Our results revealed that pig herds showed slight or no seroconversion after three to four weeks post-PCV2 immunization. The presence of PCV2 was observed during the window period for seroconversion in all herds. In conclusion, natural exposure of PCV2 occurs in the growing to fattening period, and viremia can last until slaughter. Additionally, our findings indicate that using ELISA showed the level of antibodies and aided in the understanding and surveillance of the current PCV2 status in the field.

Immunoenhancement of Recombinant Neisseria meningitides PorB Protein on Porcine Circovirus Type 2 and Mycoplasma hyopneumoniae Genetically Engineered Vaccines

2019 ◽  
Vol 26 (10) ◽  
pp. 776-784
Author(s):  
Rui Yang ◽  
Yu Tao ◽  
Gaojian Li ◽  
Jian Chen ◽  
Jianhong Shu ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Mycoplasma Hyopneumoniae ◽  
Porcine Circovirus Type ◽  
Porcine Circovirus Type 2 ◽  
Genetically Engineered ◽  
Porcine Circovirus ◽  
Practical Applications ◽  
Neisseria Meningitides ◽  
Short Period

Background:Porcine circovirus and Mycoplasma hyopneumoniae can cause respiratory diseases in pigs, which cause serious economic loss in the worldwide pig industry. Currently, these infections are mainly prevented and controlled by vaccination. The new vaccines on the market are mainly composed of subunits and inactivated vaccines but usually have lower antigenicity than traditional live vaccines. Thus, there is an increasing need to develop new adjuvants that can cause rapid and long-lasting immunity to enhance the antigenic efficacy for vaccines. Studies have shown that meningococcal porin PorB can act as a ligand to combine with Toll-like receptors to activate the production of immunological projections and act as a vaccine immunological adjuvant.Objective:In this article, we expressed and purified the recombinant PorB protein and verified its immunogenicity against porcine circovirus type 2 and Mycoplasma hyopneumoniae genetically engineered vaccine.Methods:In this article, we used prokaryotic expression to express and purify recombinant PorB protein, four different concentrations of PorB protein, Freund's adjuvant with two genetically engineered vaccines were combined with subcutaneous immunization of mice.Results:Our study shows that the appropriate dose of the recombinant protein PorB can enhance the levels of humoral and cellular responses induced by two genetically engineered vaccines in a short period of time in mice. The PorB adjuvant group may cause statistically higher antibody titers for both genetically engineered vaccines compared to Freund's commercial adjuvant (P<0.001).Conclusion:The recombinant protein PorB may be a good candidate adjuvant for improving the protective effect of vaccines against porcine circovirus type 2 and Mycoplasma hyopneumoniae, and the protein can be used for future practical applications.

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