Diagnosis of tick-borne encephalitis by a mu-capture immunoglobulin M-enzyme immunoassay based on secreted recombinant antigen produced in insect cells.
Acute tick-borne encephalitis is diagnosed by detection of IgM antibodies to tick-borne encephalitis virus (TBEV) (genus Flavivirus) in patient serum. TBEV membrane (M) and envelope (E) proteins have previously been shown to form virus-like particles when expressed in mammalian cells. We expressed the prM/M and E proteins in insect cells with a recombinant baculovirus system and obtained antigenic protein secreted into the cell culture medium, as evidenced by detection by a panel of five monoclonal antibodies to TBEV E protein. According to the sedimentation pattern in sucrose gradient centrifugation, the proteins were most likely secreted as virus-like particles. A mu-capture immunoglobulin M-enzyme immunoassay (IgM-EIA) test was developed and compared to a commercially available TBEV-IgM test (Progen) based on inactivated purified TBEVs. With a panel of 100 TBEV-IgM-negative, 50 TBEV-IgM-positive, and seven dengue virus-IgM-positive serum samples from our diagnostic laboratory, a sensitivity of 100% and specificity of 99% were obtained, and the correlation coefficient of EIA absorbances with the reference test was 0.93. The antigen was also suitable for IgG antibody detection in an immunofluorescent assay format. This is the first time that secreted, fully antigenic E protein has been produced in insect cells for this arthropod-borne flavivirus.
Jääskeläinen
Anu
A
Department of Virology, Haartman Institute, Faculty of Veterinary Medicine, FIN-00014 University of Helsinki, Finland. anu.jaaskelainen@helsinki.fi
Han
Xiuqi
X
Niedrig
Matthias
M
Vaheri
Antti
A
Vapalahti
Olli
O
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000276:immunology*; D016367:Baculoviridae / Q000235:genetics; D002499:Centrifugation, Density Gradient; D004669:Encephalitis Viruses, Tick-Borne / Q000276:immunology*; D004675:Encephalitis, Tick-Borne / Q000175:diagnosis*; D005455:Fluorescent Antibody Technique; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000097:blood; D007075:Immunoglobulin M / Q000097:blood*; D011994:Recombinant Proteins / Q000276:immunology; D018411:Spodoptera
D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D011994:Recombinant Proteins