Figure 3. Cholesterol-enriched lipid raft microdomains are important for ZEBOV entry.

(A) ZEBO-VLPs associate with lipid rafts. Vero cells were incubated with gfpZEBO-VLP (green) at 37°C for 15 min and unbound virus was removed by washing. Lipid rafts were visualized by first incubating the cells with Alexafluor₅₉₄-labeled CTxB (red) followed by coalescing the small raft domains with anti-CTxB antibody. The samples were then fixed and images taken by confocal microscopy. A mid z-section of the cells is shown. Insets i and ii are enlarged images of the indicated areas. (B) Cholesterol sequestering drugs inhibit ZEBOV infection. Vero cells were pretreated with the indicated concentrations of methyl-β cyclodextrin or nystatin for 1 h. Cells were then washed extensively to remove the drugs, and gfpZEBOV was added at an MOI of 0.1. After 24 h, cells were washed and fixed. Images were then taken with a 10× objective lens. The number of foci of infected (gfp-expressing) cells were counted for 4 images per sample in duplicate. The average number of foci is indicated ± st.dev. Similar results were obtained with HEK293T cells (not shown).