SARS-CoV-2 ZsGreen Virus Particle Assay

VeroE6 cells were transfected with pCC1-4K-SARS-CoV-2-ZsGreen plasmid using Lipofectamine 3000 (Invitrogen) and ZsGreen-expressing viral particles were harvested form culture media, as described by Rihn et al. (2021) (link). VeroE6/TMPRSS2 cells were maintained in DMEM with 200 nM glutamine, 5% FBS and 200 μg/ml Hygromycin-B (Gibco) until seeding at 1.5 × 10⁵ cells/well. After 24 h, cells were transfected with ASOs as described above and infected with SARS-CoV-2/ZsGreen particles at MOI = 0.1 for 24 h. Cells were then detached with 0.05% trypsin (Gibco), and fixed in 4% formaldehyde solution for 30 min. Fluorescence was measured in a TECAN Infinite^® 200 PRO plate reader (Tecan Life Sciences, Männedorf, Switzerland) under 480 nm excitation and 530 nm emission in black bottom 96 well plates (Corning).