Interferon-? Inhibits Ebola Virus Infection

Rhein, Bethany A.; Powers, Linda S.; Rogers, Kai; Anantpadma, Manu; Singh, Brajesh K.; Sakurai, Yasuteru; Bair, Thomas; Miller-Hunt, Catherine; Sinn, Patrick; Davey, Robert A.; Monick, Martha M.; Maury, Wendy

EBOV. BALB/c peritoneal macrophages were isolated by lavage as described above and plated in the presence of 50 ng/mL murine GM-CSF or M-CSF. For prophylactic treatment, cells were treated with 20 ng/mL of murine IFNγ and/or murine TNFα for 24 hours prior to infection. Cells were refreshed with media without the addition of cytokines and transferred to a BSL4 facility at TBRI at 72 hours after cell isolation and challenged with replication-competent EBOV encoding GFP [79] at an MOI of 0.05. Cells were analyzed 24 hours following infection as described previously [25] . Briefly, the cells were fixed and stained with Hoescht 33342 dye and imaged with a Nikon Ti-Eclipse microscope using the high-content-analysis software package (Nikon, Tokyo, Japan). Images were processed and nuclei identified by Hoescht dye fluorescence. Total cells and cells expressing GFP were then counted using Cell Profiler software (Broad Institute, MIT, Boston, Massachusetts) using customized pipelines that are available from R. Davey upon request. Data are expressed as percent GFP positive cells.