BD PharmingenTM PE Rat Anti-Mouse IL-6

Description

Expression of IL-6 by stimulated BALB/c cells. BALB/c bone marrow cells were cultured for 2 weeks in the presence of recombinant mouse GM-CSF 20 ng/ml final concentration; Cat. No. 554586 . The cells were washed and stimulated with recombinant mouse IFN-gamma 10 ng/ml final concentration; Cat. No. 554587 for 4 hours followed by an overnight stimulation with LPS 1 mug/ml final concentration; Sigma, Cat. #L-8272 and GolgiPlug 1 mug/ml final concentration; Cat. No. 555029; aka Brefeldin A . Adherent cells were incubated with 1x trypsin EDTA at 37C for 10 minutes and gently dislodged by pipetting for cell harvest. Nonspecific binding of cell surface antigens was blocked by incubation of the cells with Fc Block 10 mug/ml final concentration; Cat. No. 553142 . The cells were fixed , permeabilized then subsequently stained with 0.06 mug of PE-rat anti-mouse IL-6 antibody PE-MP5-20F3, Cat. No. 554401 by using Pharmingens staining protocol left panel . To demonstrate specificity of staining, the binding by the PE-MP5-20F3 antibody was blocked by preincubation of the PE-MP5-20F3 antibody with recombinant mouse IL-6 0.12 mug; Cat. No. 554582; right panel and by preincubation of the fixed/permeabilized cells with unlabeled MP5-20F3 antibody 5 mug; Cat. No. 554400; right panel prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabeled antibody blocking specificity controls. Expression of IL-6 by stimulated BALB/c cells. BALB/c bone marrow cells were cultured for 2 weeks in the presence of recombinant mouse GM-CSF 20 ng/ml final concentration; Cat. No. 554586 . The cells were washed and stimulated with recombinant mouse IFN-gamma 10 ng/ml final concentration; Cat. No. 554587 for 4 hours followed by an overnight stimulation with LPS 1 mug/ml final concentration; Sigma, Cat. #L-8272 and GolgiPlug 1 mug/ml final concentration; Cat. No. 555029; aka Brefeldin A . Adherent cells were incubated with 1x trypsin EDTA at 37C for 10 minutes and gently dislodged by pipetting for cell harvest. Nonspecific binding of cell surface antigens was blocked by incubation of the cells with Fc Block 10 mug/ml final concentration; Cat. No. 553142 . The cells were fixed , permeabilized then subsequently stained with 0.06 mug of PE-rat anti-mouse IL-6 antibody PE-MP5-20F3, Cat. No. 554401 by using Pharmingens staining protocol left panel . To demonstrate specificity of staining, the binding by the PE-MP5-20F3 antibody was blocked by preincubation of the PE-MP5-20F3 antibody with recombinant mouse IL-6 0.12 mug; Cat. No. 554582; right panel and by preincubation of the fixed/permeabilized cells with unlabeled MP5-20F3 antibody 5 mug; Cat. No. 554400; right panel prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabeled antibody blocking specificity controls. Expression of IL-6 by stimulated BALB/c cells. BALB/c bone marrow cells were cultured for 2 weeks in the presence of recombinant mouse GM-CSF 20 ng/ml final concentration; Cat. No. 554586 . The cells were washed and stimulated with recombinant mouse IFN-gamma 10 ng/ml final concentration; Cat. No. 554587 for 4 hours followed by an overnight stimulation with LPS 1 mug/ml final concentration; Sigma, Cat. #L-8272 and GolgiPlug 1 mug/ml final concentration; Cat. No. 555029; aka Brefeldin A . Adherent cells were incubated with 1x trypsin EDTA at 37C for 10 minutes and gently dislodged by pipetting for cell harvest. Nonspecific binding of cell surface antigens was blocked by incubation of the cells with Fc Block 10 mug/ml final concentration; Cat. No. 553142 . The cells were fixed , permeabilized then subsequently stained with 0.06 mug of PE-rat anti-mouse IL-6 antibody PE-MP5-20F3, Cat. No. 554401 by using Pharmingens staining protocol left panel . To demonstrate specificity of staining, the binding by the PE-MP5-20F3 antibody was blocked by preincubation of the PE-MP5-20F3 antibody with recombinant mouse IL-6 0.12 mug; Cat. No. 554582; right panel and by preincubation of the fixed/permeabilized cells with unlabeled MP5-20F3 antibody 5 mug; Cat. No. 554400; right panel prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabeled antibody blocking specificity controls. Show More Expression of IL-6 by stimulated BALB/c cells. BALB/c bone marrow cells were cultured for 2 weeks in the presence of recombinant mouse GM-CSF 20 ng/ml final concentration; Cat. No. 554586 . The cells were washed and stimulated with recombinant mouse IFN-gamma 10 ng/ml final concentration; Cat. No. 554587 for 4 hours followed by an overnight stimulation with LPS 1 mug/ml final concentration; Sigma, Cat. #L-8272 and GolgiPlug 1 mug/ml final concentration; Cat. No. 555029; aka Brefeldin A . Adherent cells were incubated with 1x trypsin EDTA at 37C for 10 minutes and gently dislodged by pipetting for cell harvest. Nonspecific binding of cell surface antigens was blocked by incubation of the cells with Fc Block 10 mug/ml final concentration; Cat. No. 553142 . The cells were fixed , permeabilized then subsequently stained with 0.06 mug of PE-rat anti-mouse IL-6 antibody PE-MP5-20F3, Cat. No. 554401 by using Pharmingens staining protocol left panel . To demonstrate specificity of staining, the binding by the PE-MP5-20F3 antibody was blocked by preincubation of the PE-MP5-20F3 antibody with recombinant mouse IL-6 0.12 mug; Cat. No. 554582; right panel and by preincubation of the fixed/permeabilized cells with unlabeled MP5-20F3 antibody 5 mug; Cat. No. 554400; right panel prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabeled antibody blocking specificity controls. Show More Expression of IL-6 by stimulated BALB/c cells. BALB/c bone marrow cells were cultured for 2 weeks in the presence of recombinant mouse GM-CSF 20 ng/ml final concentration; Cat. No. 554586 . The cells were washed and stimulated with recombinant mouse IFN-gamma 10 ng/ml final concentration; Cat. No. 554587 for 4 hours followed by an overnight stimulation with LPS 1 mug/ml final concentration; Sigma, Cat. #L-8272 and GolgiPlug 1 mug/ml final concentration; Cat. No. 555029; aka Brefeldin A . Adherent cells were incubated with 1x trypsin EDTA at 37C for 10 minutes and gently dislodged by pipetting for cell harvest. Nonspecific binding of cell surface antigens was blocked by incubation of the cells with Fc Block 10 mug/ml final concentration; Cat. No. 553142 . The cells were fixed , permeabilized then subsequently stained with 0.06 mug of PE-rat anti-mouse IL-6 antibody PE-MP5-20F3, Cat. No. 554401 by using Pharmingens staining protocol left panel . To demonstrate specificity of staining, the binding by the PE-MP5-20F3 antibody was blocked by preincubation of the PE-MP5-20F3 antibody with recombinant mouse IL-6 0.12 mug; Cat. No. 554582; right panel and by preincubation of the fixed/permeabilized cells with unlabeled MP5-20F3 antibody 5 mug; Cat. No. 554400; right panel prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabeled antibody blocking specificity controls. Expression of IL-6 by stimulated BALB/c cells. BALB/c bone marrow cells were cultured for 2 weeks in the presence of recombinant mouse GM-CSF 20 ng/ml final concentration; Cat. No. 554586 . The cells were washed and stimulated with recombinant mouse IFN-gamma 10 ng/ml final concentration; Cat. No. 554587 for 4 hours followed by an overnight stimulation with LPS 1 mug/ml final concentration; Sigma, Cat. #L-8272 and GolgiPlug 1 mug/ml final concentration; Cat. No. 555029; aka Brefeldin A . Adherent cells were incubated with 1x trypsin EDTA at 37C for 10 minutes and gently dislodged by pipetting for cell harvest. Nonspecific binding of cell surface antigens was blocked by incubation of the cells with Fc Block 10 mug/ml final concentration; Cat. No. 553142 . The cells were fixed , permeabilized then subsequently stained with 0.06 mug of PE-rat anti-mouse IL-6 antibody PE-MP5-20F3, Cat. No. 554401 by using Pharmingens staining protocol left panel . To demonstrate specificity of staining, the binding by the PE-MP5-20F3 antibody was blocked by preincubation of the PE-MP5-20F3 antibody with recombinant mouse IL-6 0.12 mug; Cat. No. 554582; right panel and by preincubation of the fixed/permeabilized cells with unlabeled MP5-20F3 antibody 5 mug; Cat. No. 554400; right panel prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the recombinant cytokine blocking and unlabeled antibody blocking specificity controls.