Concentration of the Causative Agent in the Filtrate of the Rous Chicken Sarcoma.

The studies of Lewis and Andervont, 1 and those of Michaelis, 2 also of Baker and Macintosh, 3 Frankel 4 and Sugiura and Benedict, 5 within the past 2 or 3 years, have disclosed additional information concerning the physical and chemical properties of the causative agent of the Rous Chicken Sarcoma. Most of these observers investigated the range of H ion concentration in which the active agent retains its infectivity, and concluded that the nature of the buffer solution appears to be of no great significance, as long as the pH is between 3.8 and 10 to 11. Since the experiments of Lewis and Michaelis were carried out with tumor extracts, and those of Sugiura and Benedict with fragments of tumor tissue, we were interested in determining the activity of the causative agent in a cell-free filtrate under varying pH values, and to correlate this with the results obtained with the tumor extract, or tissue fragments.

We found that buffering the active filtrate with a potassium hydrogen pthalate solution to pH 4 caused the solution to become cloudy, with subsequent precipitation upon standing. Upon centrifuging the suspension for about 30 minutes, a heavy precipitate was formed, and the supernatant fluid became clear. The clear supernatant fluid when tested by inoculation into chickens, is non-infective. Only when it remains slightly cloudy, indicating that all of the protein substances have not been carried down in the precipitate, will an occasional tumor result after a much delayed period of time. The precipitated portion while still at pH 4 when injected into chickens is only slightly infective, that is, its infectivity seems somewhat reduced. In other words, the active agent is carried down in the precipitated portion, and rendered partly inactive at this H ion concentration.