2.7. Plaque Reduction Assay

Confluent monolayers of MDCK cells were infected with influenza virus A/WSN/1933 (H1N1) at 37°C for 2 h. After incubation, the cell monolayer was covered with the overlay medium containing LBO (50 μg/mL and 100 μg/mL) and further cultured at 37°C in 5% CO₂ for 72 h. Subsequently, the overlay medium was removed, and the cell monolayer was fixed with 4% paraformaldehyde and stained with 1% crystal violet, and the plaques were visualized with ImmunoSpot S6 (CTL, Ohio, USA).