A Method for the Purification of the Bacteriophage.

Abstract

Numerous observers have reported the adsorption of bacteriophage corpuscles by both electro-positive and electro-negative colloids. D'Herelle 1 in a review of the literature concludes that in an alkaline medium the bacteriophage is adsorbed only by electro-positive colloids and therefore carries a negative charge, a conclusion in agreement with cataphoresis studies.

De Necker 2 reported the partial flocculation of the bacteriophage by aluminum hydroxide, an electro-positive sol. When the alumina mass is dissolved by the careful addition of acetic acid the bacteriophage reappears in the liquid. Sommer, Sommer, and Meyer 3 reported the isolation of botulinus toxin by selective adsorption on colloidal aluminum hydroxide, elution with secondary ammonium phosphate, removal of salts by dialysis, and evaporation to dryness at 40° C. Their method was used in the following attempt to purify the bacteriophage.

When a suspension of the staphylococcus bacteriophage in Martin's broth (pH 7.8) is stirred with an equal volume of a 3.5% suspension of aluminum hydroxide for one hour the bacteriophage is completely removed from the mother liquor. After washing the precipitate 4 to 6 times with distilled water no bacteriophage corpuscles can be detected in the final wash water. When the precipitate is triturated with a 0.5% solution of secondary ammonium phosphate (pH 8.0) the bacteriophage is liberated and can be separated from the alumina by centrifugation or filtration.

The final procedure adopted is as follows: 50 cc. of a suspension of the staphylococcus bacteriophage in Martin's broth is stirred for one hour with an equal volume of the aluminum hydroxide suspension. This is collected on a Buchner funnel and washed 5 times with distilled water.