A versatile flow cytometry-based immunofluorescence inhibition assay for the detection of bovine viral diarrhea virus-specific antibodies.
A fast, sensitive and reliable flow cytometry-based (FACS = fluorescence activated cell sorting) immunofluorescence inhibition assay (FACS-IFI) for the detection of virus-specific antibodies in sera is described. The method was evaluated using sera from cattle experimentally infected with bovine viral diarrhea virus (BVDV). Virus-infected cells, which were fixed and permeabilized, were incubated with diluted sera from immunized or control animals. Monoclonal antibodies (mabs) against different viral proteins were added, and detected with ALEXA488-conjugated goat-antimouse antibodies. The fluorescence signals were detected by flow cytometry and determined as mean channel values. Results were expressed as percent fluorescence inhibition compared to standardized negative sera. The FACS-IFI test with sera from experimentally infected animals was highly sensitive and specific. Comparison of the FACS-IFI results with a commercially available blocking ELISA, an indirect ELISA and the standard serum neutralization test showed a strong correlation. Furthermore, the detection of protein-specific antibodies was possible using the FACS-IFI test.
Beer
M
M
Institute of Diagnostic Virology, Friedrich-Loeffler-Institut, Greifswald-lnsel Riems, Germany. Martin.Beer@fli.bund.de
Poll
G
G
Wolf
G
G
Kaaden
O R
OR
D000818:Animals; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral / Q000097:blood*; D001912:Bovine Virus Diarrhea-Mucosal Disease / Q000175:diagnosis*; D002417:Cattle; D001908:Diarrhea Viruses, Bovine Viral / Q000276:immunology* / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary; D005434:Flow Cytometry / Q000379:methods / Q000662:veterinary; D019084:Fluorescent Antibody Technique, Indirect / Q000379:methods / Q000662:veterinary; D009500:Neutralization Tests / Q000662:veterinary; D012680:Sensitivity and Specificity; D016036:Seroepidemiologic Studies
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral