During the titration of encephalomyocarditis virus on primary mouse cells it was observed that both the size and number of plaques were increased by pretreatment of the cells with saponin which was being used to lyse erythrocytes.

Crude brown saponin was freshly dissolved for each experiment to give a 2% solution in 50% ethanol. Saponin from batch 205470/630707 prepared by British Drug Houses Ltd was used for all experiments. Cells were obtained by trypsinization (Dulbecco, 1952) of the embryos from 17-day pregnant Porton white mice. Before centrifugation the trypsinized cells were exposed for 5 min. at room temperature to 0.02% saponin. They were then centrifuged and grown as monolayers in 10% calf serum in medium 199. The cultures were infected with Habel’s large plaque (τ) mutant of encephalomyocarditis virus grown in mouse brain and were incubated in 5% CO under an overlay of medium 199 containing 0.176% bicarbonate, 5% calf serum, 5% chick embryo extract and 0.75% Difco Noble agar.


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