Serological reactivity of baculovirus-expressed Ebola virus VP35 and nucleoproteins.
Ebola virus (EBOV) is a member of the family Filoviridae and is classified as a biosafety level 4 virus. This classification makes the preparation of antigen and performance of diagnostic assays time-consuming and complicated. The objective of this study was to evaluate the value of EBOV immunoassays based on recombinant nucleoprotein (r-NP) and recombinant VP35 (r-VP35) using large serum panels of African origin and from primates. Furthermore, we investigated whether the results obtained with EBOV r-VP35 enzyme-linked immunosorbent assay (ELISA) could improve on the findings obtained with the EBOV r-NP ELISA. The full-length EBOV NP and VP35 of the EBOV subtype Zaire were expressed as histidine-tagged recombinant proteins in the baculovirus expression system. The antigenic reactivity and specificity of these recombinant proteins were determined by Western blotting and ELISA using EBOV specific monoclonal antibodies. The results obtained with the r-NP and r-VP35 ELISAs were compared with the results obtained in an indirect immunofluorescence assay based on native EBOV subtype Zaire. EBOV specific monoclonal antibodies reacted specifically with the respective proteins in both Western blot and ELISA. Five hundred and twenty six samples from humans and primates were tested with r-NP and r-VP35 ELISAs. Monkey serum samples positive for EBOV subtype Reston and Zaire were both positive in the EBOV r-NP ELISA, whereas only the EBOV Zaire infected monkeys were positive in the r-VP35 ELISA. The sensitivity and specificity values of the EBOV recombinants' ELISAs compared to those of the immunofluorescence assay were 92% and 99% for r-NP and 44% and 100% for r-VP35. r-NP ELISA proved to be a sensitive and specific assay for EBOV diagnosis and for epidemiological studies for both EBOV subtypes Reston and Zaire. The use of r-VP35 in an ELISA format has no additional value for EBOV serodiagnosis.
Groen
Jan
J
Institute of Virology, Erasmus MC, Dr. Molenwaterplein 40, 3015GD, Rotterdam, The Netherlands.
van den Hoogen
Bernadette G
BG
Burghoorn-Maas
Chantal P
CP
Fooks
Anthony R
AR
Burton
Jane
J
Clegg
Chris J S C
CJ
Zeller
Herve
H
Osterhaus
Albert D M E
AD
D000293:Adolescent; D000328:Adult; D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D016367:Baculoviridae / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D029043:Ebolavirus / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D019084:Fluorescent Antibody Technique, Indirect; D019142:Hemorrhagic Fever, Ebola / Q000276:immunology / Q000662:veterinary / Q000821:virology; D006801:Humans; D008251:Macaca; D008875:Middle Aged; D008992:Monkey Diseases / Q000276:immunology / Q000821:virology; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D011994:Recombinant Proteins / Q000276:immunology; D012680:Sensitivity and Specificity; D014758:Viral Core Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
D000914:Antibodies, Viral; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D011994:Recombinant Proteins; D014758:Viral Core Proteins; C084818:nucleoprotein VP35, Ebola virus