BD PharmingenTM FITC Rat Anti-Mouse IL-10

Description

Expression of IL-10 by stimulated CD4+ Balb/c spleen cells. Purified splenic CD4+ cells from 6 month old BALB/c mice were stimulated with plate-bound anti-CD3 145-2C11, Cat. No. 553057 at 25 mug/ml and soluble anti-mouse CD28 clone 37.51, Cat. No. 553294 at 2 mug/ml for 2 days in culture together with recombinant mouse IL-2 10 ng/ml, Cat. No. 550069 and recombinant mouse IL-4 1 ng/ml, Cat. No. 550067 , followed by a 3 day incubation with only recombinant IL-2 and IL-4. This was followed by a 5 hour stimulation with plate-bound anti-CD3 25 mug/ml and anti-mouse CD28 2 mug/ml in the presence of 2 muM monensin BD GolgiStop Cat. No. 554724 . The cells were harvested, stained with PE Rat anti-Mouse CD4 RM4-5, Cat. No. 553049 , fixed, permeabilized, and subsequently stained with FITC Rat anti-Mouse IL-10 JES5-16E3, Cat. No. 562037 by using the BD Pharmingen staining protocol. To demonstrate specificity of staining, the binding of FITC-JES5-16E3 was blocked by the preincubation of the conjugated antibody with molar excess of recombinant mouse IL-10 Cat. No. 550070; middle panel , and by preincubation of the fixed/permeabilized cells with an excess of the unlabelled JES5-16E3 antibody Cat. No. 554464; right panel . The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with the recombinant cytokine blocking middle panel and unlabelled antibody blocking right panel specificity controls. A suitable rat IgG2b isotype control for assessing background staining on fixed/permeabilized mouse cells is FITC-A95-1 Cat. No. 556923 ; use at comparable concentrations to antibody of interest. Expression of IL-10 by stimulated CD4+ Balb/c spleen cells. Purified splenic CD4+ cells from 6 month old BALB/c mice were stimulated with plate-bound anti-CD3 145-2C11, Cat. No. 553057 at 25 mug/ml and soluble anti-mouse CD28 clone 37.51, Cat. No. 553294 at 2 mug/ml for 2 days in culture together with recombinant mouse IL-2 10 ng/ml, Cat. No. 550069 and recombinant mouse IL-4 1 ng/ml, Cat. No. 550067 , followed by a 3 day incubation with only recombinant IL-2 and IL-4. This was followed by a 5 hour stimulation with plate-bound anti-CD3 25 mug/ml and anti-mouse CD28 2 mug/ml in the presence of 2 muM monensin BD GolgiStop Cat. No. 554724 . The cells were harvested, stained with PE Rat anti-Mouse CD4 RM4-5, Cat. No. 553049 , fixed, permeabilized, and subsequently stained with FITC Rat anti-Mouse IL-10 JES5-16E3, Cat. No. 562037 by using the BD Pharmingen staining protocol. To demonstrate specificity of staining, the binding of FITC-JES5-16E3 was blocked by the preincubation of the conjugated antibody with molar excess of recombinant mouse IL-10 Cat. No. 550070; middle panel , and by preincubation of the fixed/permeabilized cells with an excess of the unlabelled JES5-16E3 antibody Cat. No. 554464; right panel . The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with the recombinant cytokine blocking middle panel and unlabelled antibody blocking right panel specificity controls. A suitable rat IgG2b isotype control for assessing background staining on fixed/permeabilized mouse cells is FITC-A95-1 Cat. No. 556923 ; use at comparable concentrations to antibody of interest. Expression of IL-10 by stimulated CD4+ Balb/c spleen cells. Purified splenic CD4+ cells from 6 month old BALB/c mice were stimulated with plate-bound anti-CD3 145-2C11, Cat. No. 553057 at 25 mug/ml and soluble anti-mouse CD28 clone 37.51, Cat. No. 553294 at 2 mug/ml for 2 days in culture together with recombinant mouse IL-2 10 ng/ml, Cat. No. 550069 and recombinant mouse IL-4 1 ng/ml, Cat. No. 550067 , followed by a 3 day incubation with only recombinant IL-2 and IL-4. This was followed by a 5 hour stimulation with plate-bound anti-CD3 25 mug/ml and anti-mouse CD28 2 mug/ml in the presence of 2 muM monensin BD GolgiStop Cat. No. 554724 . The cells were harvested, stained with PE Rat anti-Mouse CD4 RM4-5, Cat. No. 553049 , fixed, permeabilized, and subsequently stained with FITC Rat anti-Mouse IL-10 JES5-16E3, Cat. No. 562037 by using the BD Pharmingen staining protocol. To demonstrate specificity of staining, the binding of FITC-JES5-16E3 was blocked by the preincubation of the conjugated antibody with molar excess of recombinant mouse IL-10 Cat. No. 550070; middle panel , and by preincubation of the fixed/permeabilized cells with an excess of the unlabelled JES5-16E3 antibody Cat. No. 554464; right panel . The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with the recombinant cytokine blocking middle panel and unlabelled antibody blocking right panel specificity controls. A suitable rat IgG2b isotype control for assessing background staining on fixed/permeabilized mouse cells is FITC-A95-1 Cat. No. 556923 ; use at comparable concentrations to antibody of interest. Show More Expression of IL-10 by stimulated CD4+ Balb/c spleen cells. Purified splenic CD4+ cells from 6 month old BALB/c mice were stimulated with plate-bound anti-CD3 145-2C11, Cat. No. 553057 at 25 mug/ml and soluble anti-mouse CD28 clone 37.51, Cat. No. 553294 at 2 mug/ml for 2 days in culture together with recombinant mouse IL-2 10 ng/ml, Cat. No. 550069 and recombinant mouse IL-4 1 ng/ml, Cat. No. 550067 , followed by a 3 day incubation with only recombinant IL-2 and IL-4. This was followed by a 5 hour stimulation with plate-bound anti-CD3 25 mug/ml and anti-mouse CD28 2 mug/ml in the presence of 2 muM monensin BD GolgiStop Cat. No. 554724 . The cells were harvested, stained with PE Rat anti-Mouse CD4 RM4-5, Cat. No. 553049 , fixed, permeabilized, and subsequently stained with FITC Rat anti-Mouse IL-10 JES5-16E3, Cat. No. 562037 by using the BD Pharmingen staining protocol. To demonstrate specificity of staining, the binding of FITC-JES5-16E3 was blocked by the preincubation of the conjugated antibody with molar excess of recombinant mouse IL-10 Cat. No. 550070; middle panel , and by preincubation of the fixed/permeabilized cells with an excess of the unlabelled JES5-16E3 antibody Cat. No. 554464; right panel . The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with the recombinant cytokine blocking middle panel and unlabelled antibody blocking right panel specificity controls. A suitable rat IgG2b isotype control for assessing background staining on fixed/permeabilized mouse cells is FITC-A95-1 Cat. No. 556923 ; use at comparable concentrations to antibody of interest. Show More Expression of IL-10 by stimulated CD4+ Balb/c spleen cells. Purified splenic CD4+ cells from 6 month old BALB/c mice were stimulated with plate-bound anti-CD3 145-2C11, Cat. No. 553057 at 25 mug/ml and soluble anti-mouse CD28 clone 37.51, Cat. No. 553294 at 2 mug/ml for 2 days in culture together with recombinant mouse IL-2 10 ng/ml, Cat. No. 550069 and recombinant mouse IL-4 1 ng/ml, Cat. No. 550067 , followed by a 3 day incubation with only recombinant IL-2 and IL-4. This was followed by a 5 hour stimulation with plate-bound anti-CD3 25 mug/ml and anti-mouse CD28 2 mug/ml in the presence of 2 muM monensin BD GolgiStop Cat. No. 554724 . The cells were harvested, stained with PE Rat anti-Mouse CD4 RM4-5, Cat. No. 553049 , fixed, permeabilized, and subsequently stained with FITC Rat anti-Mouse IL-10 JES5-16E3, Cat. No. 562037 by using the BD Pharmingen staining protocol. To demonstrate specificity of staining, the binding of FITC-JES5-16E3 was blocked by the preincubation of the conjugated antibody with molar excess of recombinant mouse IL-10 Cat. No. 550070; middle panel , and by preincubation of the fixed/permeabilized cells with an excess of the unlabelled JES5-16E3 antibody Cat. No. 554464; right panel . The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with the recombinant cytokine blocking middle panel and unlabelled antibody blocking right panel specificity controls. A suitable rat IgG2b isotype control for assessing background staining on fixed/permeabilized mouse cells is FITC-A95-1 Cat. No. 556923 ; use at comparable concentrations to antibody of interest. Expression of IL-10 by stimulated CD4+ Balb/c spleen cells. Purified splenic CD4+ cells from 6 month old BALB/c mice were stimulated with plate-bound anti-CD3 145-2C11, Cat. No. 553057 at 25 mug/ml and soluble anti-mouse CD28 clone 37.51, Cat. No. 553294 at 2 mug/ml for 2 days in culture together with recombinant mouse IL-2 10 ng/ml, Cat. No. 550069 and recombinant mouse IL-4 1 ng/ml, Cat. No. 550067 , followed by a 3 day incubation with only recombinant IL-2 and IL-4. This was followed by a 5 hour stimulation with plate-bound anti-CD3 25 mug/ml and anti-mouse CD28 2 mug/ml in the presence of 2 muM monensin BD GolgiStop Cat. No. 554724 . The cells were harvested, stained with PE Rat anti-Mouse CD4 RM4-5, Cat. No. 553049 , fixed, permeabilized, and subsequently stained with FITC Rat anti-Mouse IL-10 JES5-16E3, Cat. No. 562037 by using the BD Pharmingen staining protocol. To demonstrate specificity of staining, the binding of FITC-JES5-16E3 was blocked by the preincubation of the conjugated antibody with molar excess of recombinant mouse IL-10 Cat. No. 550070; middle panel , and by preincubation of the fixed/permeabilized cells with an excess of the unlabelled JES5-16E3 antibody Cat. No. 554464; right panel . The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with the recombinant cytokine blocking middle panel and unlabelled antibody blocking right panel specificity controls. A suitable rat IgG2b isotype control for assessing background staining on fixed/permeabilized mouse cells is FITC-A95-1 Cat. No. 556923 ; use at comparable concentrations to antibody of interest.