Early serodiagnosis of acute human cytomegalovirus infection by enzyme-linked immunosorbent assay using recombinant antigens.
DNA fragments from eight different reading frames of human cytomegalovirus (HCMV) were generated by PCR and subsequently cloned and expressed in Escherichia coli in fusion with glutathione S-transferase. The recombinant viral antigens were evaluated in immunoblot analyses. The most reactive antigens were purified and further evaluated in ELISAs. For this, sera from healthy blood donors and immunocompetent individuals with acute HCMV infection, and follow-up sera from transplant recipients with acute primary HCMV infection were used. The results of our experiments indicate that only three particular recombinant polypeptides from two viral proteins are necessary for serodiagnosis. While a fragment covering amino acids (aa) 495 to 691 of pp150 (150/1) was the most suitable antigen for the identification of infected individuals in general, immunoglobulin M antibodies against the C-terminal parts of pp150 (aa 862 to 1048; 150/7) and p52 (aa 297 to 433; 52/3) proved to be excellent serological markers to monitor acute HCMV infection. The selected recombinant antigens enable the improvement of serodiagnosis of HCMV-related diseases, especially during the early stages of infection.
Research Department, Biotest AG, Dreieich.
R
R
Vornhagen
B
B
Plachter
W
W
Hinderer
T H
TH
The
J
J
Van Zanten
L
L
Matter
C A
CA
Schmidt
H H
HH
Sonneborn
G
G
Jahn
D000914:Antibodies, Viral / Q000097:blood; D000956:Antigens, Viral / Q000235:genetics; D003001:Cloning, Molecular; D003587:Cytomegalovirus / Q000235:genetics / Q000276:immunology; D003586:Cytomegalovirus Infections / Q000175:diagnosis* / Q000209:etiology / Q000276:immunology; D004274:DNA, Recombinant / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D005069:Evaluation Studies as Topic; D005982:Glutathione Transferase / Q000235:genetics; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007075:Immunoglobulin M / Q000097:blood; D016030:Kidney Transplantation / Q000009:adverse effects; D016364:Reading Frames; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D012698:Serologic Tests / Q000379:methods*
D000914:Antibodies, Viral; D000956:Antigens, Viral; D004274:DNA, Recombinant; D004279:DNA, Viral; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase