Studies on the antigenic specificity of Mycobacterium leprae. II. Immunological characterization.

Antigens from untreated and autoclaved Mycobacterium leprae obtained through chromatofocusing were tested for their ability to both induce as well as elicit skin reactivity in guinea pigs sensitized either with homologous and heterologous mycobacteria or with the fractions derived from autoclaved M. leprae. In former studies, of the several antigen-positive fractions, one showed specific activity and the remaining others cross-reactivity, as indicated by studies of hypersensitivity. The fraction exhibiting specificity contained only one antigen, whereas the other fractions contained more than one antigen. Because of the technique employed, all the fractions contained antigens that were proteinic in nature. In the latter studies, the antigens obtained from autoclaved M. leprae were shown to possess sensitizing capabilities, in addition to induction properties. Two fractions from untreated M. leprae had antigens that were heat-labile, whereas the remaining fractions contained heat-stable antigens.

Studies on the antigenic specificity of Mycobacterium leprae. III. Further studies on immunological characterization.

The ability of the various protein antigens of Mycobacterium leprae to induce as well as detect delayed type hypersensitivity has been confirmed by studies in mice. Additionally, one of the fractions obtained from untreated M. leprae has been shown to possess specificity to the organism through immuno-analysis, thus confirming previous observations on skin-reactivity in guinea pigs. SDS-PAG electrophoresis has shown that this fraction contains a single antigen. A suggestion has been made that this single protein could be a target antigen for early diagnosis of leprosy, specifically in contacts of leprosy patients. It could also assist in detecting a latent infection. Additional studies, using different parameters, should lead to further confirmation of its specificity. It has also been suggested that such M. leprae-specific protein antigens could play an important role in the immune response of leprosy patients. They could also have a significant impact as possible immuno-protective agents, either by themselves or in combination with other immuno-potentiating agents.

Studies on the antigenic specificity of Mycobacterium leprae. I. Isoelectric and chromatofocusing separation.

Cell sonicates of Mycobacterium leprae and other mycobacteria were subjected to isoelectric focusing and chromatofocusing to evaluate their protein antigens and to determine if the patterns were significantly different. Isoelectric focusing showed that the proteins of all mycobacteria focused within the pH range of 3.5 to 5.5, except those of M. leprae which extended beyond 5.5 to 6.5. These studies have indicated for the first time that the protein antigens of mycobacteria are acidic in nature. Comparison between the proteins of untreated and autoclaved M. leprae showed distinct differences between the two preparations, in respect of loss of some antigens in the autoclaved M. leprae sonicate. This indicates that the bands that were not visible in the autoclaved M. leprae were those of heat-labile proteins. It is possible, however, that the absent bands could have been of a low order of intensity and hence were not discernible. On the other hand, the proteins could have coagulated due to the heat treatment, thus causing confirmational changes or ionic interactions with membrane components, due to their acidic nature. It is possible that the proteins in the autoclaved M. leprae are the ones that possess immunogenic properties since the protective ability of heat-killed M. leprae has already been established. Chromatofocusing studies have confirmed the isoelectric focusing data in respect of the number of antigens and their respective protein content, besides permitting the availability of the various fractions for further biological characterization.