Ebola virus neutralizing assay

Vero E6 cells were cultured in each well of a 96-well tissue culture plate. The following day, when the cells were approximately 80% confluent, 100 µg/mL of each of the 5 antibodies, as well as 2G4 used as a positive control for neutralization [6], were serially diluted in cDMEM. To this, an equal volume of Ebola virus Makona (GenBank No. {"type":"entrez-nucleotide","attrs":{"text":"KJ660348.2","term_id":"674810554","term_text":"KJ660348.2"}}KJ660348.2) was added so that each well would receive 100 plaque forming units. The virus-antibody mixture was incubated for 1 hour and then 100 µL of the virus-antibody mixture was added to the 96-well. After incubating the 96-well plate for 1 hour at 37℃, the mixture was removed and 200 µL of 2% FBS DMEM was added to each well. The plate was returned to the 37℃ incubator and plates were read for cytopathic effect 13-day post-infection. This assay was performed in the biosafety level 4 laboratory at the National Microbiology Laboratory, part of the public Health Agency of Canada in Winnipeg, Manitoba, Canada.