Recombinant VP7-based enzyme-linked immunosorbent assay for detection of immunoglobulin G antibodies to Colorado tick fever virus.
VP6, VP7, VP9, VP10, VP11, and VP12 of Colorado tick fever virus (CTF virus), a virus member of the genus Coltivirus, family Reoviridae, were expressed in bacteria with the pGEX-4T-2 vector. A partial sequence of VP7 (designated pVP7) was chosen to elaborate an enzyme-linked immunosorbent assay (ELISA) for detecting anti-CTF virus immunoglobulin G (IgG) antibodies in humans. This was based on two observations: (i) among all expressed proteins, pVP7 showed the highest immunoreactivity to an anti-CTF virus hyperimmune ascitic fluid; (ii) to provide the highest selectivity of antibody detection, the expressed sequence was chosen within a region which is highly divergent (49% amino acid identity) from the homologous sequence of another coltivirus, the Eyach virus. The pVP7 ELISA was evaluated with 368 serum samples from French blood donors and found to provide 98.1% specificity. Assays with the Calisher set of human serum samples, positive for anti-CTF virus antibodies (C. H. Calisher, J. D. Poland, S. B. Calisher, and L. A Warmoth, J. Clin. Microbiol. 22:84-88, 1985), showed that the pVP7 ELISA provided 100% sensitivity for the tested population. After elaboration of recombinant-protein-based ELISAs for diagnosis of infections with members of the viral genera Orbivirus, Orthoreovirus, and Rotavirus, it was shown that a recombinant protein could be used to detect antibodies to the human pathogen Colorado tick fever virus.
Mohd Jaafar
Fauziah
F
Unité des Virus Emergents, EA 3292, IFR 48, Université de la Méditerranée, Faculté de Médecine de Marseille, Marseille 13005, France.
Attoui
Houssam
H
Gallian
Pierre
P
Biagini
Philippe
P
Cantaloube
Jean-François
JF
de Micco
Philippe
P
de Lamballerie
Xavier
X
D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000235:genetics; D001483:Base Sequence; D036022:Capsid Proteins / Q000235:genetics / Q000276:immunology*; D003121:Colorado Tick Fever / Q000276:immunology / Q000821:virology; D003122:Colorado tick fever virus / Q000235:genetics / Q000276:immunology*; D004279:DNA, Viral / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000706:statistics & numerical data; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity
D000914:Antibodies, Viral; D000956:Antigens, Viral; D036022:Capsid Proteins; D004279:DNA, Viral; D007074:Immunoglobulin G; D011994:Recombinant Proteins