Fluorescence-based antiviral assay for the evaluation of compounds against vaccinia virus, varicella zoster virus and human cytomegalovirus.
Recombinant vaccinia virus (VACV), varicella zoster virus (VZV) and two human cytomegaloviruses (HCMV) expressing the green fluorescent protein (GFP) and the enhanced yellow fluorescent protein (EYFP) were used to develop a fluorescence-based assay for testing antiviral compounds. Infection of human embryonic lung fibroblasts (HEL) with the different recombinant viruses produced stable and detectable amount of GFP and EYFP signal as quantitated by automated fluorometry. The sensitivity of the recombinant viruses to a panel of antiviral drugs was measured and the fluorescence-based assay was compared to the cytopathic effect reduction assay (CPE-RA) in case of VACV and HCMV or to the plaque reduction assay (PRA) in case of VZV. The 50% inhibitory concentration (IC(50)) values for reference anti-pox and anti-herpesvirus compounds were comparable to those determined by CPE-RA or PRA assays. Furthermore the fluorimetric data could be confirmed by a flow cytometry assay. GFP- and EYFP-recombinant viruses proved to be a convenient tool for the evaluation of antiviral agents.
Dal Pozzo
Fabiana
F
Rega Institute for Medical Research, K.U. Leuven, Minderbroedersstraat 10, 3000 Leuven, Belgium. fabiana.dalpozzo@libero.it
Andrei
Graciela
G
Daelemans
Dirk
D
Winkler
Michael
M
Piette
Jacques
J
De Clercq
Erik
E
Snoeck
Robert
R
D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D003587:Cytomegalovirus / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D014645:Herpesvirus 3, Human / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D006801:Humans; D008826:Microbial Sensitivity Tests / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods*; D011995:Recombination, Genetic; D015203:Reproducibility of Results; D014616:Vaccinia virus / Q000187:drug effects* / Q000235:genetics / Q000502:physiology
D000998:Antiviral Agents; D049452:Green Fluorescent Proteins