J Vet Sci.  2017 Sep;18(3):307-316. 10.4142/jvs.2017.18.3.307.

Development of an immunochromatographic strip for detection of antibodies against porcine reproductive and respiratory syndrome virus

• Li H
• Yang J
• Bao D
• Hou J
• Zhi Y
• Yang Y
• Ji P
• Zhou E
• Qiao S
• Zhang G

• KMID: 2412448
• DOI: http://doi.org/10.4142/jvs.2017.18.3.307

Abstract

A simple and rapid immunochromatographic test strip incorporating a colloidal gold-labeled recombinant Nsp7 antigen probe was successfully developed for the detection of anti-porcine reproductive and respiratory syndrome virus (PRRSV) antibodies in swine. Recombinant Nsp7 protein of PRRSV labeled with colloidal gold was dispensed on a conjugate pad for use as the detector. Staphylococcal protein A and purified porcine anti-Nsp7 antibodies were blotted on a nitrocellulose membrane to form test and control lines, respectively. A comparison of the strip with standard diagnostic tests, enzyme-linked immunosorbent assays and immunoperoxidase monolayer assay, was also performed. The immunochromatographic test strip was shown to be of high specificity and sensitivity. Furthermore, the strip assay is rapid and easy to perform with no requirement for professional-level skills or equipment. It is suggested that the immunochromatographic test strip can be used to quickly and accurately detect PRRSV antibody and to be suitable for diagnostic purposes in the field.

Keyword

enzyme-linked immunosorbent assay; immunochromatography; porcine reproductive and respiratory syndrome

MeSH Terms

Animals
Antibodies, Viral/*immunology
Blotting, Western/veterinary
Enzyme-Linked Immunosorbent Assay/veterinary
Immunochromatography/veterinary
Porcine Reproductive and Respiratory Syndrome/diagnosis/immunology
Porcine respiratory and reproductive syndrome virus/*immunology
*Reagent Strips/therapeutic use
Swine
Antibodies, Viral
Reagent Strips

Figure

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  Fig. 1 Recombinant porcine reproductive and respiratory syndrome virus (PRRSV) Nsp7 preparations in Escherichia coli analyzed by SDS-PAGE and Western blot. M, protein marker; lane 1, supernatant protein of pET 28a-Nsp7 after inducing for 8h; lanes 2, soluble protein purified by Ni-NTA His Band Resin column; and lane 3, soluble protein purified by Western blot.

• 

  Fig. 2 The different assembling structure proteins identified on SDS-PAGE gels after separation by Superdex 200 gel filtration chromatography (inset). M, the protein molecular mass standard; lane 0, the total protein after purification by Ni-NTA His Band Resin; lanes 1–3, correspond to peaks 1–3 represent the larger aggregate, the dimer, and the monomer. Peak 4 was the elution of imidazole.

• 

  Fig. 3 The specific immunogenicity of recombinant Nsp7 tested by enzyme-linked immunosorbent assay. (A) Immunoreaction of the recombinant Nsp7 purified through Ni-chelating affinity chromatography with positive sera to porcine reproductive and respiratory syndrome virus (PRRSV) strain BJ-4 and HN07-1. (B–D) Immunoreaction of larger aggregate, dimer, and monomer separated by a gel filtration chromatography with positive sera to PRRSV strain BJ-4 and HN07-1.

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  Fig. 4 Specificity of the immunochromatographic strip test. Positive sera of porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus-1 (PCV-1), pseudorabies virus (PRV), classical swine fever virus (CSFV), porcine parvovirus (PPV), porcine circovirus-2 (PCV-2), and PRRSV-negative sera were tested by using the immunochromatographic strip at the same time. Result positivity can be decided by visual judgment.

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