Expression and purification of an Epstein-Barr virus encoded 23-kDa protein and characterization of its immunological properties.
Serodiagnosis of Epstein-Barr virus (EBV) infection is currently based on the detection of antibodies to distinct EBV antigens by immunofluorescence and enzyme-linked immunosorbent assay-based tests, or in part on the detection of heterophile antibodies by the Paul-Bunnell-Davidson heterophile assay. In the past few years, the specificity and the sensitivity of serodiagnostic assay systems has been improved considerably by the use of purified recombinant EBV antigens. Screening of EBV-positive sera for antigenic reactivities by immunoprecipitation with extracts of EBV-positive cells revealed a 23-kDa protein (p23) that was recognized by antibodies from all EBV carriers tested. Open reading frame BLRF2 was identified as the coding region for this protein. After cloning and high-level expression of the BLRF2 open reading frame as DHFR fusion protein in Escherichia coli, the recombinant protein was purified to near homogeneity with the help of continuous elution electrophoresis. Sera from both EBV-positive and -negative donors were screened by immunoblot analysis and enzyme-linked immunosorbent assay for IgM and IgG antibodies against the EBV-encoded protein p23. Since anti-p23 antibodies were not detectable in 30 of 30 EBV-negative sera, and 294 of 302 EBV-positive sera had either IgM and/or IgG antibody responses to this protein, recombinant p23 seems to be a useful diagnostic marker for EBV-infection.
Reischl
U
U
Institute for Medical Microbiology and Hygiene, University of Regensburg, Germany.
Gerdes
C
C
Motz
M
M
Wolf
H
H
D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D000956:Antigens, Viral / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D017931:DNA Primers; D004926:Escherichia coli; D015870:Gene Expression; D004854:Herpesvirus 4, Human / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D011237:Predictive Value of Tests; D011993:Recombinant Fusion Proteins / Q000235:genetics; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification
D000914:Antibodies, Viral; D000956:Antigens, Viral; D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D013762:Tetrahydrofolate Dehydrogenase