For virus neutralization test, serial two-fold dilutions of sera were prepared in duplicate in a 96-well tissue culture plate in 50 ml cell media (Minimal Essential Medium containing Earle's salts and supplemented with 2 mM glutamine, antibiotic-antimycotic and 10% fetal calf serum). An equal volume containing 200 TCID 50 of target virus was added and the virus-sera mix incubated for 30 min at 37uC in a humidified 5% CO 2 incubator. 100 ml of Vero cell suspension containing 2610 5 cells/ml was added and the plate incubated at 37uC in a humidified 5% CO 2 incubator. After 4 days, the plate was examined for viral CPE. The highest serum dilution generating complete inhibition of CPE is defined as the final neutralizing titer.