Renilla luciferase rluc assay system

A library of 1,058 botanical drug compounds was purchased from Weikeqi Biotech (Sichuan, China). The compounds were stored as 10 mM stock solutions until use. HTS was carried out as shown in Fig. 1A. Vero cells were treated in duplicate with the compounds (50 μM); 1 h later, cells were infected with LASVrv (MOI = 0.1) for 1 h. After 23 h, cells were fixed with 4% paraformaldehyde and stained with 4′,6-diamidino-2-phenylindole (DAPI; Sigma-Aldrich, St. Louis, MO, USA). Nine fields per well were imaged on an Operetta high-content imaging system (PerkinElmer, Waltham, MA, USA), and the percentages of infected and DAPI-positive cells were calculated using Harmony 3.5 (PerkinElmer). Cell viability was evaluated using a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. To determine the IC₅₀, LASVpv at an MOI of 0.1 was used utilizing the timeline described above. Luciferase activity was measured using the Renilla luciferase (Rluc) assay system (Promega, Madison, WI, USA), and IC₅₀ was calculated using GraphPad Prism 6.

A library of 1015 fragment-based drugs was purchased from Selleck Chemicals (Cat: L1600; Houston, TX, USA). Compounds were stored in 10 mM stock solution in DMSO at −80 °C until use. HTS was carried out as shown in Figure 1A. Vero cells were treated in duplicate with the compounds (100 μM); 1 h later, cells were infected with LASV_VSVpv (MOI, 0.05) for 1 h. After 23 h, the infected cells were lysed, and luciferase activity was measured using the Renilla luciferase (Rluc) assay system (Promega, Madison, WI, USA). Cell viability was evaluated by using the Cell Counting Kit-8 (CCK-8) assay. In order to determine the IC₅₀, LASV_VSVpv at an MOI of 0.05 was used utilizing the timeline described above. Luciferase activity was measured using the Rluc assay system, and IC₅₀ was calculated using GraphPad Prism 8.