Methodologies currently used in
our laboratory in the study of Aspergillus fumigatus:
Fungal cultures in fermentor under controled conditions.
Molecular biology techniques
Cloning and disruption of genes by integration
of auxotrophic or resistance markers.
Expression of A. fumigatus proteins
in Pichia pastoris.
Transcriptome analysis using macroarrays containing
5,000 PCR products.
Biochemistry and immunology techniques
Extraction and chromatographic
purification of peptides, proteins and polysaccharides (liquid chromatography,
Localization, identification, caracterization of proteins by biochemical
and immunochemical methods.
Classical techniques of carbohydrate analysis
(GC, GC/MS, NMR).
Production of antibodies directed against fungal proteins
Cellular biology techniques
Cultures of alveolar macrophages (primary
human and murine cells, cell line MHS),
Study of phagocytosis (internalization,
intracellular trafic, killing of fungal conidia) by immunofluorescence (double
Study of virulence of strains of Aspergillus
fumigatus in mice (intranasal inoculations),
Production of antibodies
in rabbit or mouse (intranasal, intradermal, intraveinous, intramuscular inoculations
of proteinic ou polysaccharidic antigens or DNA).
Ultrastructure and immunocytochimistry in transmission and scanning electron microscopy