Inhibition assay on viral replication

The viral replication inhibition assay was according to that described previously [20]. 100 μL of 100pfu PrV were incubated with the monolayer Vero cells in a 24-well flat-bottom plate under 5 % CO₂ at 37 °C for 90 min. The incubating solution was discarded, the cells treated with 0.5 mL of either 25, 50, 100, or 200 μg/mL extract in 1 % methylcellulose and reincubated under 5 % CO₂ at 37 °C for 48 h. The infected cells were fixed with 10 % formal saline and stained with 0.5 % crystal violet solution. The plaques were counted and the IC₅₀ calculated as described in plaque reduction assay.