BD Pharmingen™ FITC Mouse IgG1, κ Isotype Control
Clone MOPC-21 (also known as MOPC21; MOPC 21) (RUO)
Expression of IFN-γ by stimulated human peripheral blood mononuclear cells (PBMC). Human PBMC were stimulated for 6 hours with PMA (50 ng/ml final concentration; Sigma, Cat. No. P-8139) and calcium ionophone A23187 (250 ng/ml final concentration; Sigma, Cat. No. C-9275) in the presence of GolgiStop™ (2 µM final concentration; Cat. No. 554724). The PBMC were stained with PE-Cy5 (formerly referred to as BD-Cy-Chrome™) - anti-CD3 (Cy-Chrome™ - UCHT1, Cat. No. 555334), fixed, permeabilized, and subsequently stained with 0.25 µg of FITC-mouse anti-human IFN-γ antibody (FITC-4S.B3, Cat. No. 554551; left panel) or FITC-MOPC-21 immunoglobulin (0.25 µg; Cat. No. 554679; right panel) by using the BD Bioscience staining protocol. To demonstrate specificity of staining, the binding of FITC-4S.B3 was blocked by preincubation of fixed/permeabilized cells with excess unlabelled 4S.B3 antibody (5 µg; Cat. No. 554549). The quadrant markers for the bivariate dot plot were set based on autofluorescence controls and verified using the unlabelled 4S.B3 antibody blocking control.
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Expression of IFN-γ by stimulated human peripheral blood mononuclear cells (PBMC). Human PBMC were stimulated for 6 hours with PMA (50 ng/ml final concentration; Sigma, Cat. No. P-8139) and calcium ionophone A23187 (250 ng/ml final concentration; Sigma, Cat. No. C-9275) in the presence of GolgiStop™ (2 µM final concentration; Cat. No. 554724). The PBMC were stained with PE-Cy5 (formerly referred to as BD-Cy-Chrome™) - anti-CD3 (Cy-Chrome™ - UCHT1, Cat. No. 555334), fixed, permeabilized, and subsequently stained with 0.25 µg of FITC-mouse anti-human IFN-γ antibody (FITC-4S.B3, Cat. No. 554551; left panel) or FITC-MOPC-21 immunoglobulin (0.25 µg; Cat. No. 554679; right panel) by using the BD Bioscience staining protocol. To demonstrate specificity of staining, the binding of FITC-4S.B3 was blocked by preincubation of fixed/permeabilized cells with excess unlabelled 4S.B3 antibody (5 µg; Cat. No. 554549). The quadrant markers for the bivariate dot plot were set based on autofluorescence controls and verified using the unlabelled 4S.B3 antibody blocking control.
Preparation And Storage
Recommended Assay Procedures
Immunofluorescent Staining and Flow Cytometric Analysis: The FITC-MOPC-21 immunoglobulin (Cat. No. 554679) is a suitable mouse IgG1 isotype control for assessing the level of background staining on paraformaldehyde-fixed/saponin-permeabilized mouse or human cells for flow cytometric analysis. Use at comparable concentrations to antibody of interest (e.g., ≤ 0.5 µg mAb/1 million cells) (figure right panel).
Product Notices
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
Companion Products
The MOPC-21 immunoglobulin is a mouse myeloma protein. The MOPC-21 immunoglobulin was selected as an isotype control following screening for low background on a variety of mouse and human tissues.
This antibody is routinely tested by flow cytometric analysis. Other applications were tested at BD Biosciences Pharmingen during antibody development only or reported in the literature.
Development References (1)
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Prussin C, Metcalfe DD. Detection of intracytoplasmic cytokine using flow cytometry and directly conjugated anti-cytokine antibodies. J Immunol Methods. 1995; 188(1):117-128. (Biology: Flow cytometry). View Reference
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.