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Separation of Tetanus Antitoxin Dissociated from a Toxin-Antitoxin Precipitate¹

From the Division of Bacteriology, Department of Biological Sciences, Purdue University, Lafayette, Indiana

Abstract

The foregoing study has shown that after treatment with pepsin of a toxin-antitoxin precipitate dissolved at pH 3, a substantial quantity of inactive precipitate may be removed by adjusting the pH to about 4.7 and heating in a water bath of 45°C for a short time. After removing the precipitate, the clear supernatant, which contained free antitoxin, was concentrated and fractionated at varying concentrations of ammonium sulfate. Most of the antitoxin activity was precipitable at between 40 and 50% saturation with ammonium sulfate. The fraction obtained under the latter conditions showed only one electrophoretic component, when studied in a phosphate buffer of 0.20 ionic strength and pH 7.7 at a protein concentration of 1%. It had an antitoxic activity of 1,470 units per mg nitrogen by the flocculation test and 1,330 units per mg nitrogen by the animal protection tests.

Footnotes

¹ Part of a thesis submitted to the faculty of Purdue University by A. E. Bolyn in partial fulfulfillment of the requirements for the degree of Doctor of Philosophy.

² Present address: National Drug Company. Biological Division, Swiftwater, Pennsylvania.