Further Observations on the Use of Pneumococcus Extracts in Effecting Transformation of Type in Vitro

Pneumococcus extracts highly active in inducing the in vitro transformation of the specific types of Pneumococcus have been prepared by dissolving S cells with sodium desoxycholate, precipitating the dissolved material in alcohol in which the bile salt remains soluble, and extracting the precipitate in salt solution. Further purification of these active extracts has been attained by the removal of considerable inactive material by charcoal adsorption and by reprecipitation of the adsorbed extract in alcohol or acetone. The importance of using young cultures for extraction, and of preventing autolysis during the preparation of the extracts, is emphasized. Extracts prepared by the method described have been filtered through Berkefeld Candles (V, N, and W) without appreciable loss in activity, provided the reaction of the extract was slightly alkaline at the time of filtration. The purified and filtered extracts are water-clear, and sterile by rigid cultural and animal tests. They have been heated to temperatures of 60 degrees C. for 30 minutes without appreciable loss in their capacity to induce specific changes in type. And although they have generally shown definite decrease in potency after heating to temperatures above 80 degrees C., some extracts have been found active even after an exposure of 10 minutes to a temperature of 90 degrees C. They have been completely inactivated by boiling. Relatively small amounts of extract have been effective when added to a broth medium containing normal serum or serous fluid. In this medium, R pneumococci, irrespective of their type derivation, have developed and thereafter retained all the type-specific characteristics of the encapsulated S cells from which the extract was prepared. The specific action of the extracts is discussed with reference to their transforming and antigenic properties.