The Purification and Concentration of Diphtheria Toxoid

Abstract

By precipitation with acetone at 4°C. a refined diphtheria toxoid was obtained as a dry powder, readily soluble in aqueous solutions. The powder itself appears to be stable. When dissolved in half the original volume of physiological salt solution, the toxoid remained stable, in the cold room, for a period of 7 months. Only toxoids should be used which have been completely detoxified by the treatment with formalin. The formalin is immediately and completely removed by precipitation with acetone. Filtration through a Mandler filter or the addition of a preservative did not appear to have any deleterious effect. Incubating at 39°C. and freezing for 20 hours did not impair its activities. The flocculating value of the purified toxoid containing phenol was almost completely destroyed by freezing, while when merthiolate was the preservative, it was unchanged. Approximately 62 per cent of the nitrogen was eliminated in the process with approximately 15 per cent loss in flocculating value. The flocculating values were nearly double those of the original material and thus corresponded with the concentration, which was onehalf the volume. By this method, the antigenic activity of a toxoid below standard may be increased to a satisfactory standard. The antigenic activity of the refined product in the immunization of guinea pigs may in general parallel its flocculating values but the relation between the results of these tests may not be proportional nor constant. As an immunizing agent in the guinea pig, the refined toxoid possesses an antigenic activity that is equal to, or greater than, that of the unconcentrated toxoid.