Optimal Conditions of Plaque Titration of Japanese Encephalitis Virus on BHK21 Cells
摘要
Optimal conditions for the infectivity titration by plaque formation (PFU) of Japanese encephalitis (JE) virus on mammalian BHK21 cells were investigated. Early virus inoculation at 6 hr after seeding low concentration of the cells (0.25×10^5/ml) produced larger and clearer plaques compared with higher concentration or longer incubation (1-2 days) of the cells. However, the virus titer increased by the incubation time until 2 days to become the highest for this inoculum cell concentration, and up to 1 day for the inoculum cell concentration of 0.5 or 1.0×10^5/ml. While the virus titer did not differ significantly by the time of cell culture for the inoculum cell concentration of 2.0×10^5/ml. Supplementation of 9% fetal bovine serum (FBS) to the cell growth medium was mandatory for clear plaque formation. Among several methyl cellulose (MC) concentrations in the overlay medium, a moderate concentration of 1.25% was optimal to produce clear and distinct plaques on 1 day's culture from 2.0×10^5 cells/ml.