Expression of the nucleoprotein gene of rabies virus for use as a diagnostic reagent.
The nucleoprotein (N) gene of rabies virus CTN strain, was cloned, sequenced and expressed in Escherichia coli as a fusion with maltose binding protein (MBP). The antigenicity of this recombinant MBP-N fusion protein was examined by Western blotting and enzyme linked immunosorbent assay (ELISA). Subsequently, an indirect ELISA was developed to detect rabies specific antibody levels. Using sera from naive and vaccinated animals the ELISA results were compared with virus neutralizing antibodies detected by a rapid fluorescent focus inhibition test (RFFIT). Neutralizing titres by RFFIT were found to correlate well with the OD values in the ELISA (r=0.9436) and the sensitivity and specificity of the ELISA were shown to be 93.4 and 100%, respectively. The data indicate that the recombinant MBP-N fusion protein can be expressed and isolated straightforwardly and may be useful as a safe and abundant source of antigen to monitor seropositivity in vaccinated canines.
He
Y
Y
Laboratory for Animal Molecular Virology, College of Biological Sciences, China Agricultural University, Beijing 100094, China.
Gao
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D
Zhang
M
M
D000818:Animals; D000835:Animals, Wild; D000914:Antibodies, Viral / Q000097:blood*; D015153:Blotting, Western; D003001:Cloning, Molecular; D004285:Dogs; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D005455:Fluorescent Antibody Technique; D051379:Mice; D009500:Neutralization Tests; D019590:Nucleocapsid Proteins / Q000235:genetics* / Q000276:immunology*; D011819:Rabies Vaccines; D011820:Rabies virus / Q000235:genetics* / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000276:immunology; D012680:Sensitivity and Specificity
D000914:Antibodies, Viral; D019590:Nucleocapsid Proteins; D011819:Rabies Vaccines; D011993:Recombinant Fusion Proteins; C099601:nucleocapsid protein, Rabies virus