Generation of Pseudotyped VSVs

Vesicular stomatitis virus (VSV) (Indiana) containing the green fluorescent protein (GFP) gene instead of the VSV glycoprotein (G) gene (VSVΔG*-G) and pseudotyped VSVs bearing GPs of LASV (strain Josiah) or LUJV (strain IGR140) were generated as described previously (15 (link), 34 (link)). Briefly, HEK293T cells were transfected with the expression plasmid pCAGGS carrying the LUJV GP gene, and 24 h later, the cells were infected with VSVΔG*-G at a multiplicity of infection of 1.0. After a 16-h incubation, the supernatants were collected and centrifuged to remove cell debris. Infectious units (IU) of the pseudotyped VSVs in each cell line were determined as described previously (35 (link), 36 (link)). Briefly, cells were seeded into 96-well plates 1 day before virus inoculation, and 24 h later, GFP-positive cells were counted using an In Cell analyzer 2000 system (GE Healthcare). To reduce the background infectivity of the residual parent VSVΔG*-G, each pseudotyped virus stock was treated with a neutralizing monoclonal antibody specific for the VSV G protein (VSV-G[N]1-9) before use (37 (link)).