Purification of Rabies Virus Grown in Tissue Culture

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Abstract
Extracellular rabies virus, grown in monolayer cultures of [baby hamster kidney] BHK21 cells in the presence of medium supplemented with bovine serum albumin, was purified by the following procedure. Virus was precipitated from infectious tissue culture fluid by zinc acetate and was resuspended in a solution of EDTA. The suspension was filtered through a Sephadex column and was treated with RNase and DNase. The virions were then pelleted by centrifugation at high speed and were resuspended in buffer solution. Banding of the virus by centrifugation in a sucrose density gradient was the final step in the purification procedure. Purified preparations contained bullet-shaped virus particles of variable length and little (up to 5%) contaminating host-cell material. Most of the virions were "complete", i.e., 180 mm long, but some virus particles were shorter. The length distribution of the virions was non-random. Shorter virions seemed to be noninfectious and showed markedly decreased hemagglutinating activity. The complement-fixing activity and the RNA to protein ratio of the virions were not related to the length of the virus particles. Although the properties of extracellular and intracellular viruses were similar, the procedure was not suitable for purification of intracellular rabies virus.