Journal: Clinical and Developmental Immunology

Article Title: Both CD4 + and CD8 + Lymphocytes Participate in the IFN- γ Response to Filamentous Hemagglutinin from Bordetella pertussis in Infants, Children, and Adults

doi: 10.1155/2012/795958

Figure Lengend Snippet: Effect of the CD4 + or CD8 + depletion on the FHA-induced IFN- γ production by PBMC from B. pertussis -infected or vaccinated children. Isolated PBMC were stimulated in vitro for 72 h with 1 μ g/mL FHA after CD4 + (a) or CD8 + (b) depletion. Results from infected children are represented by columns 1 to 5 whereas those from vaccinated children are indicated by numbers 6 to 16. Open columns represent the values obtained for the nondepleted cell suspension and grey columns those obtained after CD4 + or CD8 + depletions. The arrows indicate the samples characterized by a higher IFN- γ secretion in CD8-depleted cell suspension compared to the PBMC.

Article Snippet: After fixation and permeabilization of the cells according to the manufacturers' instructions (Lysing-Solution1 and Permeabilizing-Solution2, BD Biosciences, USA), PBMC were stained with anti-human monoclonal antibodies: anti-CD3-PerCp-Cy5.5, anti-CD4-APC-Cy7, anti-CD8-APC, and anti-IFN- γ -PE (BD Biosciences, USA).

Techniques: Infection, Isolation, In Vitro

Journal: Clinical and Developmental Immunology

Article Title: Both CD4 + and CD8 + Lymphocytes Participate in the IFN- γ Response to Filamentous Hemagglutinin from Bordetella pertussis in Infants, Children, and Adults

doi: 10.1155/2012/795958

Figure Lengend Snippet: Effect of blocking anti-MHC class I and anti-MHC class II antibodies on the FHA-specific IFN- γ production. Isolated PBMC were stimulated in vitro for 72 h with 1 μ g/mL FHA in the presence or absence of blocking anti-MHC-II (a) and (b) or anti-MHC-I (c) and (d) antibodies or in the presence or absence of control isotypes. Unstimulated cells were tested as control. IFN- γ concentrations were measured in the cell culture supernatants and values obtained for unstimulated cells were subtracted from those obtained for stimulated cells. Circles represent the individual IFN- γ concentrations obtained after stimulation with FHA in the presence of anti-MHC antibodies or control isotypes. Results obtained for the same samples are linked. * P < 0.05; ** P < 0.001; *** P < 0.0001.

Article Snippet: After fixation and permeabilization of the cells according to the manufacturers' instructions (Lysing-Solution1 and Permeabilizing-Solution2, BD Biosciences, USA), PBMC were stained with anti-human monoclonal antibodies: anti-CD3-PerCp-Cy5.5, anti-CD4-APC-Cy7, anti-CD8-APC, and anti-IFN- γ -PE (BD Biosciences, USA).

Techniques: Blocking Assay, Isolation, In Vitro, Cell Culture

Journal: Clinical and Developmental Immunology

Article Title: Both CD4 + and CD8 + Lymphocytes Participate in the IFN- γ Response to Filamentous Hemagglutinin from Bordetella pertussis in Infants, Children, and Adults

doi: 10.1155/2012/795958

Figure Lengend Snippet: Percentages and absolute numbers of FHA-induced IFN- γ producing CD4 + and CD8 + T cells in B. pertussis -infected ( n = 6) or vaccinated children ( n = 4). Isolated PBMC were stimulated in vitro for 24 h with 5 μ g/mL FHA, and IFN- γ -containing CD4 + (left columns) and CD8 + (right columns) T lymphocytes were detected by flow cytometry. (a) represents the percentages of positive cells and the absolute numbers of IFN- γ -producing cells expressed for 10 6 CD3 + T lymphocytes are shown on (b). The lines join the results obtained for the same children. Percentages obtained for unstimulated cells were subtracted from those obtained for stimulated cells. * P < 0.05, ** P < 0.001.

Article Snippet: After fixation and permeabilization of the cells according to the manufacturers' instructions (Lysing-Solution1 and Permeabilizing-Solution2, BD Biosciences, USA), PBMC were stained with anti-human monoclonal antibodies: anti-CD3-PerCp-Cy5.5, anti-CD4-APC-Cy7, anti-CD8-APC, and anti-IFN- γ -PE (BD Biosciences, USA).

Techniques: Infection, Isolation, In Vitro, Flow Cytometry

Journal: Clinical and Developmental Immunology

Article Title: Both CD4 + and CD8 + Lymphocytes Participate in the IFN- γ Response to Filamentous Hemagglutinin from Bordetella pertussis in Infants, Children, and Adults

doi: 10.1155/2012/795958

Figure Lengend Snippet: Dot-plot of the flow cytometric analyses of the phenotype of FHA-induced IFN- γ producing cells for one representative subject. Isolated PBMC were cultured in vitro for 24 h in the presence (lower panels) or absence (upper panels) of FHA. CD4 + T cells (left panels) or CD8 + T cells (right panels) were gated in the CD3 + T lymphocytes. Results are expressed as the percentages of IFN- γ producing cells among the cell subpopulation analyzed.

Article Snippet: After fixation and permeabilization of the cells according to the manufacturers' instructions (Lysing-Solution1 and Permeabilizing-Solution2, BD Biosciences, USA), PBMC were stained with anti-human monoclonal antibodies: anti-CD3-PerCp-Cy5.5, anti-CD4-APC-Cy7, anti-CD8-APC, and anti-IFN- γ -PE (BD Biosciences, USA).

Techniques: Isolation, Cell Culture, In Vitro

Journal: Clinical and Developmental Immunology

Article Title: Both CD4 + and CD8 + Lymphocytes Participate in the IFN- γ Response to Filamentous Hemagglutinin from Bordetella pertussis in Infants, Children, and Adults

doi: 10.1155/2012/795958

Figure Lengend Snippet: Effect of the CD4 + or CD8 + depletion on the FHA-induced IFN- γ production by PBMC from B. pertussis -infected or vaccinated children. Isolated PBMC were stimulated in vitro for 72 h with 1 μ g/mL FHA after CD4 + (a) or CD8 + (b) depletion. Results from infected children are represented by columns 1 to 5 whereas those from vaccinated children are indicated by numbers 6 to 16. Open columns represent the values obtained for the nondepleted cell suspension and grey columns those obtained after CD4 + or CD8 + depletions. The arrows indicate the samples characterized by a higher IFN- γ secretion in CD8-depleted cell suspension compared to the PBMC.

Article Snippet: After fixation and permeabilization of the cells according to the manufacturers' instructions (Lysing-Solution1 and Permeabilizing-Solution2, BD Biosciences, USA), PBMC were stained with anti-human monoclonal antibodies: anti-CD3-PerCp-Cy5.5, anti-CD4-APC-Cy7, anti-CD8-APC, and anti-IFN- γ -PE (BD Biosciences, USA).

Techniques: Infection, Isolation, In Vitro