BD PharmingenTM Purified Rat Anti-Mouse IL-4

Description

Immunocytochemistry analysis of IL-4 expression on mouse splenocytes. RBC-lysed BALB/c splenocytes were enriched for CD4+ cells by panning and cultured for 2 days with plate-bound Purified NA/LE Hamster Anti-Mouse CD3e Cat. No. 553057; 10 lambdag/ml for coating and soluble Purified NA/LE Hamster Anti-Mouse CD28 Cat. No. 553294; 2 lambdag/ml in the presence of Recombinant IL-2 Cat. No. 550069; 10 ng/ml and IL-4 Cat. No. 550067; 25 ng/ml . The cells were subsequently harvested, washed and recultured with mouse IL-2 and mouse IL-4 for an additional 3 days. Finally, the cells were harvested, washed and cultured with PMA Sigma, Cat. #P 8139, 5 ng/ml and ionomycin Sigma, Cat. #I-0634, 500 ng/ml in the presence of GolgiPlug Cat. No. 555029 for 4 hr at 37C. The activated cells were harvested and the presence of IL-4 producing cells was detected by immunocytochemistry using a three-step staining procedure that employs a Biotin Goat anti-Rat IgG secondary antibody Cat. No. 559286 and a horseradish peroxidase-based detection system. To demonstrate specificity of staining the binding of Purified Rat Anti-Mouse IL-4 Cat. No. 559062 antibody was blocked by the preincubation of the purified antibody with excess recombinant mouse IL-4 protein data not shown . Nomarski optics, original magnification 400 X . Immunocytochemistry analysis of IL-4 expression on mouse splenocytes. RBC-lysed BALB/c splenocytes were enriched for CD4+ cells by panning and cultured for 2 days with plate-bound Purified NA/LE Hamster Anti-Mouse CD3e Cat. No. 553057; 10 lambdag/ml for coating and soluble Purified NA/LE Hamster Anti-Mouse CD28 Cat. No. 553294; 2 lambdag/ml in the presence of Recombinant IL-2 Cat. No. 550069; 10 ng/ml and IL-4 Cat. No. 550067; 25 ng/ml . The cells were subsequently harvested, washed and recultured with mouse IL-2 and mouse IL-4 for an additional 3 days. Finally, the cells were harvested, washed and cultured with PMA Sigma, Cat. #P 8139, 5 ng/ml and ionomycin Sigma, Cat. #I-0634, 500 ng/ml in the presence of GolgiPlug Cat. No. 555029 for 4 hr at 37C. The activated cells were harvested and the presence of IL-4 producing cells was detected by immunocytochemistry using a three-step staining procedure that employs a Biotin Goat anti-Rat IgG secondary antibody Cat. No. 559286 and a horseradish peroxidase-based detection system. To demonstrate specificity of staining the binding of Purified Rat Anti-Mouse IL-4 Cat. No. 559062 antibody was blocked by the preincubation of the purified antibody with excess recombinant mouse IL-4 protein data not shown . Nomarski optics, original magnification 400 X . Show More Immunocytochemistry analysis of IL-4 expression on mouse splenocytes. RBC-lysed BALB/c splenocytes were enriched for CD4+ cells by panning and cultured for 2 days with plate-bound Purified NA/LE Hamster Anti-Mouse CD3e Cat. No. 553057; 10 lambdag/ml for coating and soluble Purified NA/LE Hamster Anti-Mouse CD28 Cat. No. 553294; 2 lambdag/ml in the presence of Recombinant IL-2 Cat. No. 550069; 10 ng/ml and IL-4 Cat. No. 550067; 25 ng/ml . The cells were subsequently harvested, washed and recultured with mouse IL-2 and mouse IL-4 for an additional 3 days. Finally, the cells were harvested, washed and cultured with PMA Sigma, Cat. #P 8139, 5 ng/ml and ionomycin Sigma, Cat. #I-0634, 500 ng/ml in the presence of GolgiPlug Cat. No. 555029 for 4 hr at 37C. The activated cells were harvested and the presence of IL-4 producing cells was detected by immunocytochemistry using a three-step staining procedure that employs a Biotin Goat anti-Rat IgG secondary antibody Cat. No. 559286 and a horseradish peroxidase-based detection system. To demonstrate specificity of staining the binding of Purified Rat Anti-Mouse IL-4 Cat. No. 559062 antibody was blocked by the preincubation of the purified antibody with excess recombinant mouse IL-4 protein data not shown . Nomarski optics, original magnification 400 X .