FIG. 3.

Bioassay for production of IFN in FEK cells infected with recombinant wild-type (rWT) and NS1 mutant viruses. (a) Schematic diagram of the bioassay for IFN production. FEK cells were infected with each of the four equine recombinant viruses. Supernatants were collected, and any virus present was UV inactivated. Fresh FEK cells were treated with the inactivated supernatants and then infected with VSV-GFP at an MOI of 0.1. (b) Time course of IFN production. Over an 11-h infection period, supernatants were taken at 2-h intervals as indicated. VSV-GFP-infected cells were examined by fluorescence microscopy, and pictures of representative fields were taken. (c) Quantification of IFN mRNA synthesis by real-time RT-PCR. Infected cells were collected at the time points indicated, and RNA was processed for real-time RT-PCR analysis for the β-actin and IFN-β genes. The results represent relative IFN-β mRNA levels normalized to levels of β-actin mRNA.