Electron Microscopy of Tissue Cultures Infected with the Virus of Eastern Equine Encephalomyelitis. ∗

Abstract

During the past 2 years we have been studying the growth of several viruses in tissue culture with particular emphasis on the use of the electron microscope. The cellular destruction and overwhelming growth of the virus of eastern equine encephalomyelitis are here reported.

Successful electron microscopy of virus infected cells has been limited to studies on “virus induced” tumors 1 , 2 and has not been applied to small rapidly multiplying viruses. Our technic is essentially that of Porter 3 except that cells are usually fixed for only ten minutes with osmic acid vapor. 4

Chick embryo tissue was cultured for 6 days with one transfer in roller tubes 5 in a completely homologous medium composed of chicken plasma and serum and chick embryo extract. Pieces of this tissue were then exposed to a 10^-2 dilution of the virus. A saline extract of embryos infected with the fourth embryo passage of a standard guinea pig passage of eastern equine encephalomyelitis was used as source of virus. This tissue explant was then placed in a special chamber. This was formed by drilling the center out of a metal plate and sandwiching it between 2 formvar coated glass slides. The medium consisted of 20% chicken serum. Within the next 24 hours of incubation at a temperature of 37°C, the cells had grown out poorly and were already beginning to be destroyed. The cells were fixed by exposure to osmic acid vapor and were either examined directly or were first coated with chromium to sharpen the image. The fluid taken from the tissue culture chamber taken just before osmic acid fixation of the preparation shows a high titer of infectivity for the chick embryo.