Plaque assay
Collecting RBCs were suspended in 1 mL PBS, then repeatedly frozen and
thawed to release the virus. Vero cells were seeded in 12-well plates
and grown to confluent monolayers. Then cells were infected with 500 μL
serial ten-fold dilutions of the frozen and thawed samples for 1 h at 37
°C. After infection, cells were overlaid with 0.7% low melting point
agarose (Solarbio) in Dulbecco’s Modified Eagle’s Medium (DMEM)
containing 2% FBS and incubated at 37
°C
for 72 h. Plaques were visualized by staining with crystal violet.