Ebola VLP-Based High-Throughput Screening

Ebola VLPs containing a beta-lactamase-fused VP40 protein (EBOV BlaVP40) and GP were produced in Dr García-Sastre's lab, as previously described.^{6 (link)} LiveBLAzer FRET–B/G Loading Kit with CCF2-AM and Opti-MEM reduced serum medium were purchased from Life Technologies (Carlsbad, CA, USA). An adenosine triphosphate (ATP) content cell viability assay kit was purchased from Promega (Madison, WI, USA). Polystyrene plates (384-well and 1536-well black, clear bottom, sterile, tissue culture treated) were purchased from Greiner Bio-One (Monroe, NC, USA). A FDA-approved drug collection of 600 compounds was originally prepared at the National Center for Advancing Translational Sciences (NCATS) for a personalized cancer treatment project. This collection excludes certain drugs, such as those known to be immunosuppressive, topically applied drugs, and those for approved use in animals. In a follow-up screening, we used an NCATS-approved drug collection of 2816 compounds that was previously assembled.⁷ All of the compounds were dissolved as a 10 mM stock solution in dimethyl sulfoxide (DMSO) and diluted in DMSO at a 1∶3 dilution to generate six concentrations in 384-well plates, followed by reformatting into three 1536-well compound source plates for HTS.