Studies of Yellow Fever Virus in Tissue Culture

Abstract

The virus of yellow fever, like most invisible agents of infectious diseases, has not been cultivated on any of the usual bacteriological media. After it had been established that this virus belongs to the group of filterable infectious agents, it seemed worth while to attempt its cultivation in vitro by methods successfully developed for other filterable viruses. A strain of yellow fever virus highly pathogenic for mice made it possible for the infectivity of cultures to be tested repeatedly and accurately on a large scale.

Of the several methods of growing viruses in tissue cultures, the use of Carrel dishes has given the best results. Only after repeated attempts were we finally able to carry the virus through the first subculture. The culture medium chiefly used has been a mixture of normal monkey serum and Tyrode solution in which were suspended small fragments of fresh tissue. Many kinds of tissue were used, principally minced kidneys and testicles of guinea pigs and rabbits or minced chicken embryos 8 to 10 days old. The minced embryos have been used almost entirely in the more recent experiments because the virus appeared to remain more uniformly virulent in the cultures containing these tissues.

The virus for the primary culture was prepared by grinding infectious mouse brain in a mortar with 9 times its weight of Tyrode solution and centrifuging the suspension for about 10 minutes at 3000 revolutions per minute. One part of the supernatant fluid was added to 4 parts of the culture medium, and the mixture was distributed in several Carrel dishes, about 2.0 cc. in each. Taking the concentration of infected mouse-brain tissue as one, the final dilution of virus would be 1:50. The cultures and subcultures were always tested for bacteriological sterility.