BD PharmingenTM FITC Rat Anti-Human IL-4

Description

Expression of IL-4 by stimulated human peripheral blood mononuclear cells PBMC . Human PBMC were stimulated with soluble anti-human CD3 antibody 1 mug/ml final concentration; Cat. No. 555329 , recombinant human IL-2 10 ng/ml final concentration; Cat. No. 554603 and recombinant human IL-4 10 ng/ml final concentration; Cat. No. 554605 for 2 days. The cells were subsequently cultured in medium containing recombinant human IL-2 and recombinant human IL-4 for 3 days. Finally, the cells were harvested and stimulated for 6 hours with PMA Sigma and calcium ionophore A23187 Sigma in the presence of GolgiStop 2muM final concentration; Cat. No. 554724 . The cells were harvested, stained with PE-Cy5-anti CD4 .25 mug final concentration; Cat. No. 555348 , fixed, permeabilized, and subsequently stained with 0.12 mug of FITC-rat anti-human IL-4 antibody FITC-MP4-25D2, Cat. No. 554484 by using the BD Pharmingen staining protocol left panel . To demonstrate specificity of staining, the binding of FITC-MP4-25D2 antibody was blocked by preincubation of the fixed/permeabilized cells with unlabelled MP4-25D2 antibody 2.5 mug, Cat. No. 554482; right panel prior to staining. The quadrant markers for the bivariate dot plot were set based on the autofluorescence controls and verified using unlabelled antibody blocking controls. Expression of IL-4 by stimulated human peripheral blood mononuclear cells PBMC . Human PBMC were stimulated with soluble anti-human CD3 antibody 1 mug/ml final concentration; Cat. No. 555329 , recombinant human IL-2 10 ng/ml final concentration; Cat. No. 554603 and recombinant human IL-4 10 ng/ml final concentration; Cat. No. 554605 for 2 days. The cells were subsequently cultured in medium containing recombinant human IL-2 and recombinant human IL-4 for 3 days. Finally, the cells were harvested and stimulated for 6 hours with PMA Sigma and calcium ionophore A23187 Sigma in the presence of GolgiStop 2muM final concentration; Cat. No. 554724 . The cells were harvested, stained with PE-Cy5-anti CD4 .25 mug final concentration; Cat. No. 555348 , fixed, permeabilized, and subsequently stained with 0.12 mug of FITC-rat anti-human IL-4 antibody FITC-MP4-25D2, Cat. No. 554484 by using the BD Pharmingen staining protocol left panel . To demonstrate specificity of staining, the binding of FITC-MP4-25D2 antibody was blocked by preincubation of the fixed/permeabilized cells with unlabelled MP4-25D2 antibody 2.5 mug, Cat. No. 554482; right panel prior to staining. The quadrant markers for the bivariate dot plot were set based on the autofluorescence controls and verified using unlabelled antibody blocking controls. Expression of IL-4 by stimulated human peripheral blood mononuclear cells PBMC . Human PBMC were stimulated with soluble anti-human CD3 antibody 1 mug/ml final concentration; Cat. No. 555329 , recombinant human IL-2 10 ng/ml final concentration; Cat. No. 554603 and recombinant human IL-4 10 ng/ml final concentration; Cat. No. 554605 for 2 days. The cells were subsequently cultured in medium containing recombinant human IL-2 and recombinant human IL-4 for 3 days. Finally, the cells were harvested and stimulated for 6 hours with PMA Sigma and calcium ionophore A23187 Sigma in the presence of GolgiStop 2muM final concentration; Cat. No. 554724 . The cells were harvested, stained with PE-Cy5-anti CD4 .25 mug final concentration; Cat. No. 555348 , fixed, permeabilized, and subsequently stained with 0.12 mug of FITC-rat anti-human IL-4 antibody FITC-MP4-25D2, Cat. No. 554484 by using the BD Pharmingen staining protocol left panel . To demonstrate specificity of staining, the binding of FITC-MP4-25D2 antibody was blocked by preincubation of the fixed/permeabilized cells with unlabelled MP4-25D2 antibody 2.5 mug, Cat. No. 554482; right panel prior to staining. The quadrant markers for the bivariate dot plot were set based on the autofluorescence controls and verified using unlabelled antibody blocking controls. Show More Expression of IL-4 by stimulated human peripheral blood mononuclear cells PBMC . Human PBMC were stimulated with soluble anti-human CD3 antibody 1 mug/ml final concentration; Cat. No. 555329 , recombinant human IL-2 10 ng/ml final concentration; Cat. No. 554603 and recombinant human IL-4 10 ng/ml final concentration; Cat. No. 554605 for 2 days. The cells were subsequently cultured in medium containing recombinant human IL-2 and recombinant human IL-4 for 3 days. Finally, the cells were harvested and stimulated for 6 hours with PMA Sigma and calcium ionophore A23187 Sigma in the presence of GolgiStop 2muM final concentration; Cat. No. 554724 . The cells were harvested, stained with PE-Cy5-anti CD4 .25 mug final concentration; Cat. No. 555348 , fixed, permeabilized, and subsequently stained with 0.12 mug of FITC-rat anti-human IL-4 antibody FITC-MP4-25D2, Cat. No. 554484 by using the BD Pharmingen staining protocol left panel . To demonstrate specificity of staining, the binding of FITC-MP4-25D2 antibody was blocked by preincubation of the fixed/permeabilized cells with unlabelled MP4-25D2 antibody 2.5 mug, Cat. No. 554482; right panel prior to staining. The quadrant markers for the bivariate dot plot were set based on the autofluorescence controls and verified using unlabelled antibody blocking controls. Show More Expression of IL-4 by stimulated human peripheral blood mononuclear cells PBMC . Human PBMC were stimulated with soluble anti-human CD3 antibody 1 mug/ml final concentration; Cat. No. 555329 , recombinant human IL-2 10 ng/ml final concentration; Cat. No. 554603 and recombinant human IL-4 10 ng/ml final concentration; Cat. No. 554605 for 2 days. The cells were subsequently cultured in medium containing recombinant human IL-2 and recombinant human IL-4 for 3 days. Finally, the cells were harvested and stimulated for 6 hours with PMA Sigma and calcium ionophore A23187 Sigma in the presence of GolgiStop 2muM final concentration; Cat. No. 554724 . The cells were harvested, stained with PE-Cy5-anti CD4 .25 mug final concentration; Cat. No. 555348 , fixed, permeabilized, and subsequently stained with 0.12 mug of FITC-rat anti-human IL-4 antibody FITC-MP4-25D2, Cat. No. 554484 by using the BD Pharmingen staining protocol left panel . To demonstrate specificity of staining, the binding of FITC-MP4-25D2 antibody was blocked by preincubation of the fixed/permeabilized cells with unlabelled MP4-25D2 antibody 2.5 mug, Cat. No. 554482; right panel prior to staining. The quadrant markers for the bivariate dot plot were set based on the autofluorescence controls and verified using unlabelled antibody blocking controls. Expression of IL-4 by stimulated human peripheral blood mononuclear cells PBMC . Human PBMC were stimulated with soluble anti-human CD3 antibody 1 mug/ml final concentration; Cat. No. 555329 , recombinant human IL-2 10 ng/ml final concentration; Cat. No. 554603 and recombinant human IL-4 10 ng/ml final concentration; Cat. No. 554605 for 2 days. The cells were subsequently cultured in medium containing recombinant human IL-2 and recombinant human IL-4 for 3 days. Finally, the cells were harvested and stimulated for 6 hours with PMA Sigma and calcium ionophore A23187 Sigma in the presence of GolgiStop 2muM final concentration; Cat. No. 554724 . The cells were harvested, stained with PE-Cy5-anti CD4 .25 mug final concentration; Cat. No. 555348 , fixed, permeabilized, and subsequently stained with 0.12 mug of FITC-rat anti-human IL-4 antibody FITC-MP4-25D2, Cat. No. 554484 by using the BD Pharmingen staining protocol left panel . To demonstrate specificity of staining, the binding of FITC-MP4-25D2 antibody was blocked by preincubation of the fixed/permeabilized cells with unlabelled MP4-25D2 antibody 2.5 mug, Cat. No. 554482; right panel prior to staining. The quadrant markers for the bivariate dot plot were set based on the autofluorescence controls and verified using unlabelled antibody blocking controls.