A lot of experimental live attenuated Japanese B encephalitis virus vaccine for human trial was prepared in 199 medium plus 2% human albumin by making a pool of repeated harvests from virus inoculated HKC monolayers at brief intervals. The titer was 7.7 log TCD50 before and 6.6 following filtration. The vaccine was free from bacteria and other detectable viral agents by all the usual safety tests.

In weanling mice by the i.c. route and in suckling mice by the s.c. route the vaccine titered less than 100. Frozen at about -70°C it has remained stable for at least 18 months. It had a conspicuous T marker, failing to grow detectably at 40°C, and also had what appeared to be a characteristic plaque form which distinguished it from wild strains. Only one monkey of those given undiluted virus i.c. died. This occurred on the 18th day. Virus isolated from the brain had partially altered characteristics. All surviving monkeys given undiluted virus by the i.c. route developed detectable neutralizing antibodies. HAI antibody response was equivocal and CF negative. Six monkeys and two chimpanzees inoculated s.c. with 1 ml of the vaccine failed to develop either fever, viremia or neutralizing antibodies. One of three of these monkeys given a second injection after 60 days developed antibodies which persisted 171 days and another responded after a third injection. Three others not given a booster were tested for viremia following an s.c. challenge with virulent virus. The viremia response appeared to be less than that of the controls. Weanling mice with reduced body temperatures proved to be somewhat more susceptible to i.c. inoculation of this virus than were those with higher temperatures and survivors were more resistant to subsequent i.c. challenge inoculation.

It is concluded that this experimental vaccine lot has all the characteristics of earlier lots made with lactalbumin hydrolysate and calf serum and that other characterization beyond that to which other lots were subjected supports further its suitability for cautious testing by the peripheral route in man. Safety appears reasonably assured.

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