Sample preparation. For infection, ,5610 8 C6/36 cells were infected with DENV at a MOI of 20 at 30uC. Infection of all cells (required for lipidomic analyses) was confirmed by immunofluorescence analyses. Each experiment included three biological replicates. Following infection, cells were harvested at 36 and 60 hr post-infection, pelleted and resuspended in 100 ml of 100 mM ammonium bicarbonate. Lipids were extracted from an equal number of cells using a modified Bligh and Dyer protocol. Briefly, a mixture of 2:1 Chloroform:methanol, 0.1% acetic acid and 0.01% butylated hydroxy toluene (BHT) were added to the cell suspension in ammonium bicarbonate such that there was a 4:1 ratio of organic solvent to cells. Following lipid extraction, the organic phase was separated from the aqueous phase by centrifugation and dried down under a N 2 stream in low retention microfuge tubes (Fisher). The dried lipids were resuspended in 75 ml of methanol and vortexed for 10 s. The samples were then centrifuged at 13,4006 g for 5 min to remove any particulates.