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Proteomics ; 1(4): 574-86, 2001 Apr.
Article in English | MEDLINE | ID: mdl-11681210

ABSTRACT

The complete sequencing of the Mycobacterium tuberculosis genome offers a unique opportunity to fully elucidate the biology of this human pathogen. One aspect of significant importance is the definition of T cell antigens. This report describes the development and implementation of a proteomic approach to defining such antigens. Large quantities of subcellular protein fractions of M. tuberculosis were resolved by two-dimensional liquid phase electrophoresis (2-D LPE), resulting in 355 and 299 fractions of culture filtrate and cytosolic proteins, respectively. Analysis of these fractions against splenocytes of C57Bl/6 mice infected with M. tuberculosis resulted in the identification of 37 fractions that stimulated a dominant T cell response, as measured by the production of interferon-gamma. Additionally, when the 2-D LPE fractions were assayed against splenocytes harvested at 10 and 40 days post infection significant changes in the T cell response were observed. Molecular characterization of the proteins contained in each of the 38 immunodominant fractions by liquid chromatography-mass spectrometry and liquid chromatography-tandem mass spectrometry resulted in the identification of 30 individual proteins. Many of these represented previously defined antigens. However 17 of these proteins were novel T cell antigens. The data presented demonstrate that proteomics offers a rapid and facile approach for elucidation of immunodominant T cell antigens of pathogenic bacteria.


Subject(s)
Antigens, Bacterial/isolation & purification , Mycobacterium tuberculosis/immunology , Proteome , T-Lymphocytes/immunology , Animals , Antigens, Bacterial/chemistry , Antigens, Bacterial/genetics , Bacterial Proteins/genetics , Bacterial Proteins/immunology , Bacterial Proteins/isolation & purification , Electrophoresis, Gel, Two-Dimensional , Genome, Bacterial , Humans , Interferon-gamma/biosynthesis , Mice , Mice, Inbred C57BL , Mycobacterium tuberculosis/genetics , Mycobacterium tuberculosis/pathogenicity , Proteome/genetics , Proteome/immunology , Proteome/isolation & purification
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