Enhanced Growth and Plaquing of Rabies Virus in Static Chick Embryo Cell Culture

Abstract

The 7-day egg passage line of HEP Flury strain of rabies virus was inoculated to primary chick embryo (CE) cells prepared in different ways to compare efficiencies of viral growth and plaguing. Special care to minimize cellular damage due to trypsin at the step of monodispersion and sowing a comparatively large number of cells for monolayer preparation were required for rabies plaguing, whereas such cares were not necessary for plaguing of vesicular stomatitis virus. Plaque number and size were increased by incorporation of a high concentration of thymidine into cell growth medium. Various other means to produce a static state of CE cells were tested, and a maximal plaguing efficiency was obtained when dishes receiving a massive number of dispersed cells in MEM plus 1% calf serum were incubated at 37 C for 1 day without any buffering for monolayer preparation and postinfection incubation was done at 32 C in a CO2-incubator.Bottle cultures of CE cells prepared in a similar manner, when infected with HEP Flury virus, yielded a markedly higher titer of virus than CE cells prepared by our previous standard method.

Alteration of the In Vitro Host Range of Rabies Virus after Serial Chick Embryo Cell Passage Using Alkaline Maintenance Medium

“…Two lines of HEP Flury strain of rabies virus [25] were compared in most experiments. One was 7-day egg passage line which represented the 283rd to 284th egg passages, and seed virus suspensions were prepared from infected embryos in the same manner as described in the accompanying paper [36]. The other line was maintained by serial passage in CE cells using maintenance medium adjusted at pH 8.2 in the manner stated earlier [44], and culture fluids representing the 46th to 55th CE passage levels were used.…”

“…PBS refers to Dulbecco and Vogt's phosphate-buffered saline free of cations [17], and BS means 0.01 M phosphatebuffered saline at pH 7.2 free of potassium. When virus titration was not performed immediately after harvesting, the specimens were stored at -70 C in the same fashion as previously stated [36]. Neutralization test.…”

Absence of Interferon Production in a Newly Established Human Cell Line

“…except for the Bl strain of NDV which did not produce clear plaques in CE cells. The CE plaque assay of rabies virus followed the method of Seklne and Yashina [3].…”