Concentration and Purification of B. Megatherium Phage

B. megatherium phage, like staphylococcus phage, is inactivated by repeated high speed centrifugation (Northrop, 1938; Hotchin, 1954). This method of purification has the additional disadvantage that large volumes of material cannot be handled. B. megatkerium phage is precipitated from peptone solution by 0.4 saturated ammonium sulfate. The precipitate is filtered off, washed, and dissolved in a small volume of 1 ~ sodium acetate. A few -/of trypsin are added and this solution kept at 0 ° for several days. The protein content of this solution is now nearly equivalent to that calculated from its active phage content. The phage may be precipitated from this solution by the slow addition (with stirring) of an equal volume of saturated magnesium sulfate. A fine silky precipitate forms which consists of highly refractile irregular particles and fibers. This precipitate contains all the phage activity. The number of plaques formed by 1 rag. nitrogen of this precipitate is very nearly equal to the number calculated from the size (Murphy, 1954) of the phage particle. If a solution of this precipitate is heated or made acid so as to inactivate the phage, the protein then appears as a flocculent amorphous precipitate. ExperimentaL--An example of the method is shown in Table I.