Factors Influencing the Inactivation of the Rabbit Papilloma Virus by X-Rays

Lacassagne found several years ago that the rabbit-papilloma virus is notably resistant to X-rays, 1 and Syverton has more recently reported that a dose of 14,000,000 r was required to abolish the infectivity of a cell-free suspension of the papilloma virus 2 though 1,000,000 r or less sufficed to inactivate certain other viruses and bacteria. In experiments undertaken lately for other purposes we have noted that the amount of irradiation required to inactivate the papilloma-virus is influenced by the virus concentration, as also by extraneous material present along with it in the irradiated suspensions. Berkefeld filtrates, which contained the virus in large amount, were rendered completely non-infectious upon exposure to 2 to 4 million r of irradiation, while only 400,000 to 800,000 r was required when the virus had been partially purified by repeated differential centrifugations.

Virus was obtained by grinding infectious cottontail rabbit papil-loma-tissue and extracting it in 10 or 20 volumes of isotonic saline and centrifuging at about 4,400 rpm for 20 minutes in an angle centrifuge, with filtration of the supernatant fluid through Berkefeld V candles. Partially purified suspensions of the virus were prepared by centrifuging the filtrate at 30,000 rpm for 1 hour in an air-driven centrifuge, after which the supernatant fluid—which contained no detectable virus but much extraneous protein, as indicated by nitrogen-determinations—was removed. The small pellet of sediment was then resuspended in isotonic saline to the original volume and spun at about 4,400 rpm for 20 minutes. The highspeed centrifugation was then repeated, the pellet of sediment re-suspended in saline, and another low-speed centrifugation done to remove gross particles. The final virus suspensions were very faintly opalescent and contained less than 1% as much nitrogen as the whole filtrate.