Thiazolyl blue tetrazolium bromide (MTT) assay

The MDCK cells were seeded on a 96-well plate at the density of 1 × 10⁴ cells/well. Y30 extract (final 0.4–25%) was prepared in water (final 0.4–25%) and mixed with infection medium [DMEM supplemented with 1% bovine serum albumin (BSA; Wako), P/S, and 4 mM l-glutamine]. The resulting mixture was added to the cells and subsequently incubated for 24 or 72 h at 37°C in the presence of 5% CO₂. After incubation, the cell viability was determined using the MTT cell proliferation assay as described previously [34,36].