Images for product: anti-Colony Stimulating Factor 2 (Granulocyte-Macrophage) (CSF2) antibody
Expression of GM-CSF by stimulated human peripheral blood mononuclear cells (PBMC). Ficoll™-separated human PBMC were stimulated with soluble anti-human CD3 antibody (1 µg/ml final concentration, UCHT1), recombinant human IL-2 (10 ng/ml final concentration) and recombinant human IL-4 (10 ng/ml final concentration) 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 (50 ng/ml final concentration, Sigma) and calcium ionophore A23187 (250 ng/ml final concentration, Sigma) in the presence of BD GolgiStop™ (2 µM final concentration). The cells were harvested, stained with PE-Cy™5 anti CD4, fixed, permeabilized, and subsequently stained with 0.25 µg of PE Rat anti-Human GM-CSF antibody (PE-BVD2-21C11, ABIN967465) by using the staining protocol (first panel). To demonstrate specificity of staining, the binding of the PE-BVD2-21C11 antibody was blocked by preincubation of the antibody conjugate with recombinant human GM-CSF (0.1 µg, Center panel) and by preincubation of the fixed/permeabilized cells with unlabeled BVD2-21C11 antibody (5 µg, ABIN967465, second panel) prior to staining. The quadrant markers for the bivariate dot plot were set based on the autofluorescence control and verified using the ligand-blocking and unlabeled antibody blocking controls.