Sars cov 2 spike neutralizing antibody

One day prior to the experiment, 1 × 104 Vero E6 cells were plated into a 96-well plate. Serum was decomplemented at 56 °C for 30 min and an initial dilution of 1:10 was prepared in OptiMEM. Two-fold dilutions were subsequently prepared and mixed with rSARS-CoV-2 NL virus (MOI = 1) for 1 h at room temperature and then plated onto cells. After a 1-h incubation at 37 °C inoculum was removed and 200 μL of fresh DMEM containing 2% FBS and 1% penicillin/streptomycin was added. After a 24 h incubation at 37 °C with 5% CO2 media was removed and cells were fixed with 10% formalin for 1 h. A SARS-CoV-2 spike neutralizing antibody (Sino Biological Inc., Beijing, China; 2 μg/μL; Clone #57) was used as a positive control. Cells were washed with 1× PBS and 20 μM furimazine (MedChem Express, Monmouth Junction, NJ, USA) luciferin substrate was added onto cells. Cells were then imaged using an IVIS spectrum imager (PerkinElmer) and analyzed using LivingImage software (PerkinElmer). Titers were determined as the reciprocal of the highest dilution with >50% reduction of cytopathic effect.