The Postendosomal Assay

BHK-21 cells were infected with LCMV (MOI = 0.1); simultaneously, to synchronize the virus in the endosome and to avoid membrane fusion, cells were also treated with 20 mM ammonium chloride for 1 h. Then the drugs were added in the presence of ammonium chloride for 30 min, after which the cells were washed to remove ammonium chloride, and virus membrane fusion started. Finally, the cells were incubated with drugs for 16 h altogether and tested by qRT-PCR. To exclude the potential effects of the drugs on replication during the postendosomal assay, a supplementary postentry assay was also developed. The operation procedures were same except being treated with ammonium chloride. BHK-21 cells were infected with LCMV (MOI = 0.1). One hour p.i., drugs were added to cells and cells were incubated at 37 °C for 16 h. Then cells were lysed and tested by qRT-PCR.