Immunofluorescence of Japanese encephalitis virus infection in vitro: localization of viral antigen in canine cerebellar tissue cultures.

Abstract

ABSTRACTPropagation of Japanese encephalitis (JE) virus in cells of dog cerebellar tissue cultures was investigated by means of fluorescent antibody (FA) technique. The fluorescent globulin conjugate was made from the serum of a dog inoculated with partially purified JE virus, treated by Sephadex G‐25 and DEAE cellulose column chromatography and then adsorbed with dog liver powder. This preparation was found to be appropriate for the present work. Fluorescence was demonstrable in virus‐infected cultures of three different types of cells, fibroblast‐like cells, nerve cells and some of the glial type cells. Fluorescence could first be demonstrated about 20 hours after virus inoculation and appeared to increase in intensity in proportion to the increase of infective virus present in the cultures. The specificity of the reaction was supported by the non‐reactivity of control (non‐infected) cultures and by the results of blocking tests. The infected nerve cells and glial type cells also exhibited morphological changes clearly detectable by the FA techniques, corresponding to the changes shown in Bodian‐stained preparations. The localization of FA antigen in the fibers of these cells suggests a possible mode of spread of JE virus in the nervous tissues. In any of the cell types studied thus far, the nuclei remained FA‐unstained even during the advanced stage of infection.