Preparation of a Highly Purified Carbohydrate-free Johnin Protein

Abstract

THE preparation of a complement-fixing antigen of carbohydrate nature from Mycobacterium johnei has been described in a previous publication¹. The present communication deals with the isolation and purification of a carbohydrate-free protein antigen from Johne's bacilli killed with phenol. This antigen has been shown to be active in complement fixation tests with sera of certain animals immunized with johnin purified protein derivative in adjuvants. Heat-killed organisms are not suitable for the preparation of the purified specific protein since disintegration products, formed at these relatively high temperatures, interact to form complexes difficult to separate. However, a ‘cold method’, based on urea extraction, proved to be practical. This relatively simple procedure yielded fair amounts of a protein devoid of serologically-active carbohydrates and of nucleic acid contaminants.