Tubercle bacillary extracts immunogenic for mice 7. electrophoretic analyses

Summary The water-soluble extract of trypsin-digested tubercle bacilli which specifically immunizes mice and guinea pigs against tuberculosis can be separated by agar gel electrophoresis into two inert fractions and one active one. The two inert constituents are cathodic and are detectable by conventional electrophoresis and immunoelectrophoresis. One is a heavy molecular weight polysaccharide, and the other appears to be a peptide. The immunizing fraction on the other hand, is strongly anodic, having a mobility 1·87 times that of human serum albumin at pH 8·2. It was detected and identified by preparatory electrophoresis coupled with in vivo tests for its immunogenicity, and its mobility then was defined with greater precision by a new technique—two-dimensional immunoelectrophoresis. This modification of conventional immunoelectrophoresis was necessary because of the extreme lability of immunoprecipitates produced by reaction between the immunogen and a specific antiserum. The rapid mobility of the immunogen also was confirmed by two-dimensional agar gel electrophoresis using 6,9-diamino-2-ethoxy-acridine as a semispecific indicator for it: this acridine dye precipitates the immunogen without precipitating the two non-immunogenic contaminants of trypsin extract. The data presented indicate that the immunogen is a very poor inducer of humoral antibody formation. Its strongly anionic charge, taken together with previously published data, suggests that is may be a polysaccharide strongly bound to a negatively charged peptide or that it may be acidic polysaccharide or mucopolysaccharide normally located in the cell wall or cell membrane of tubercle bacilli.