Purification of the 65 kD protein from Mycobacterium gordonae and use in skin test response to Mycobacterium leprae.
The cell wall-associated protein of Mycobacterium gordonae (Mr = 65,000) was purified by affinity chromatography using a murine monoclonal antibody produced in response to the crossreactive 65 kD protein of M. leprae. The affinity-purified material was analyzed for purity by protein and carbohydrate analyses, SDS-PAGE, and immunoblotting. The final preparation contained a major protein band on SDS-PAGE analysis (Mr = 65,000) with no detectable carbohydrates. The affinity fraction was prepared at 250 micrograms/ml (protein) in sterile saline and 0.1 ml injected intradermally into guinea pigs immunized 30 days earlier. Gross changes at 48 hr were consistent with the characteristics of a delayed hypersensitivity skin reaction measuring 2.5 mm, 3.4 mm, and 2.7 mm in animals which had been immunized with M. leprae, M. gordonae, or M. bovis (BCG), respectively. Histologically, all 65 kD protein skin-test sites showed marked edema and infiltration by numerous lymphocytes, macrophages, and scattered neutrophils. Animals injected with Freund's incomplete adjuvant showed a minimal or no reaction (1.8 mm) to the purified protein. These results further define the immunogenicity of the 65 kD protein of M. gordonae and by inference M. leprae, and demonstrate the ability of crossreactive epitopes of the 65 kD protein to sensitize lymphocytes involved in delayed-type hypersensitivity reaction to M. leprae, M. gordonae, and M. bovis (BCG).
D000818:Animals; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial / Q000302:isolation & purification*; D001426:Bacterial Proteins / Q000276:immunology / Q000302:isolation & purification*; D003429:Cross Reactions; D000939:Epitopes / Q000302:isolation & purification; D006168:Guinea Pigs; D051379:Mice; D009161:Mycobacterium / Q000276:immunology*; D009166:Mycobacterium leprae / Q000276:immunology*; D009170:Nontuberculous Mycobacteria / Q000276:immunology*; D012882:Skin Tests
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D000939:Epitopes