J Vet Sci.  2010 Jun;11(2):169-171. 10.4142/jvs.2010.11.2.169.

Development and characterization of stable cell lines constitutively expressing the porcine reproductive and respiratory syndrome virus nucleocapsid protein

• Sagong M
• Park CK
• Kim SH
• Moon SU
• Cho SC
• Lee C

• KMID: 1110866
• DOI: http://doi.org/10.4142/jvs.2010.11.2.169

Abstract

Despite global efforts to control porcine reproductive and respiratory syndrome virus (PRRSV) infection, the virus continues to cause economic problems in the swine industry worldwide. In this study, we attempted to generate and characterize a panel of stable BHK cell lines that constitutively express the nucleocapsid (N) protein of type 1 or type 2 PRRSV. The established BHK cell lines were found to react well with N-specific antibodies as well as the hyperimmune serum of pigs raised against each genotype of PRRSV. Taken together, the data implicate a potential usefulness for the newly generated stable cell lines as a diagnostic reagent for PRRSV serology.

Keyword

diagnostic reagent; nucleocapsid protein; PRRSV; stable BHK cell line

MeSH Terms

Animals
Antibodies, Viral/analysis/immunology
Blotting, Western/veterinary
Cell Line
Cricetinae
Female
Genotype
Nucleocapsid Proteins/genetics/*immunology
Porcine Reproductive and Respiratory Syndrome/diagnosis/*immunology
Porcine respiratory and reproductive syndrome virus/genetics/*immunology
Swine
Transfection/veterinary

Figure

• 

  Fig. 1 Constitutive expression of the genotype-specific nucleocapsid (N) protein in BHK-EU-ORF7 (A) and BHK-NA-ORF7 (B) cells. Expression levels of the N protein in individual stable cell clones were determined by western blot analysis. Each lane represents individual neomycin-resistant BHK cell clones.

• 

  Fig. 2 Immunofluorescence assay using the genotype-specific monoclonal antibodies. BHK-EU-ORF7 and BHK-NA-ORF7 cells were incubated individually with each genotype-specific antibody. A and D α-EU: EU genotype-specific monoclonal antibody (MAb), B and E α-NA: NA genotype-specific MAb, C and F α-EU + NA: genotype-common MAb. ×40.

• 

  Fig. 3 Immunoperoxidase monolayer assay. Cells were incubated with the genotype-specific MAbs or PRRSV-specific hyperimmune sera followed by staining using Vectorstain ABC peroxidase and DAB substrate kits. G and J α-LV: anti-LV pig serum, H and K α-VR-2332: anti-VR-2332 pig serum, I and L Pre-immune: normal pre-immunized pig serum. ×10.

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