参考图片
Expression of IFN-γ by stimulated CD4+ and CD4-C3H spleen cells. Splenocytes from C3H mice were stimulated in culture for 4 hours using PMA (5 ng/ml final concentration; Sigma Cat. #P-8139) and Ionomycin (500 ng/ml final concentration; Sigma Cat. #I0634) in the presence of GolgiPlug™ Protein Transport Inhibitor (1 µl/ml, Cat. No. 555029). The splenocytes were harvested and stained with 0.06 µg of FITC Rat Anti-Mouse CD4 (FITC-Rm4-5, Cat. No. 553047), fixed, permeabilized and subsequently stained with 0.12 µg of APC Rat IgG1 isotype control antibody (APC-R3-34, Cat. No. 554686, left panel) or with APC Rat Anti-Mouse IFN-γ (APC-XMG1.2, Cat. No. 554413, middle panel) by using the BD Pharmingen staining protocol. To demonstrate specificity of staining, the binding by the APC-XMG1.2 antibody was blocked by preincubation of the fixed, premeabilized cells with unlabeled XMG1.2 antibody (5.0 µg; Cat. No. 554409, right panel) prior to staining. The quadrant markers for the bivariate dot plots were set based on the autofluorescence controls and verified using the unlabeled antibody blocking specificity control.
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