PE Mouse Anti-Human IFN-γ
Clone 4S.B3 (RUO)
- Brand BD Pharmingen™
- Alternative Name IFNG; Interferon-gamma; IFG; IFI; Type II interferon
- Concentration 0.2 mg/ml
- Isotype Mouse BALB/c IgG1, κ
- Reactivity Human (QC Testing) Rhesus, Cynomolgus, Baboon (Tested in Development)
Intracellular staining (flow cytometry) (Routinely Tested)
- Immunogen Human IFN-γ from supernatants of S. aureus-stimulated PBMC
- Storage Buffer Aqueous buffered solution containing ≤0.09% sodium azide.
- Regulatory Status RUO
Regulatory Status Legend
The 4S.B3 monoclonal antibody specifically binds to interferon-γ (IFN-γ). The immunogen used to generate this hybridoma was partially purified human IFN-γ obtained from supernatants of human PBMC stimulated with Staphylococcus aureus. Interferon-γ (IFN-γ) is a potent multifunctional cytokine that is produced by several activated cell types including NK, NKT, CD4+TCRαβ+, CD8+TCRαβ+, and TCRγδ+ T cells. IFN-γ exerts its biological effects through specific binding to the high-affinity IFN-γ Receptor Complex comprised of IFN-γRα (CD119) and IFN-γRβ subunits. In addition to its antiviral effects, IFN-γ upregulates a number of lymphoid cell functions including the antimicrobial and antitumor responses of macrophages, NK cells, and neutrophils. In addition, IFN-γ can exert strong regulatory influences on the proliferation, differentiation, and effector responses of B cell and T cell subsets. These influences can involve IFN-γ's capacity to boost MHC class I and II expression by antigen-presenting cells as well as to direct effects on B cells and T cells themselves. Human IFN-γ is a 14-18 kDa glycoprotein containing 143 amino acid residues.
Clone 4S.B3 also cross-reacts with a cytoplasmic component of peripheral blood CD3+ lymphocytes of baboon, and both rhesus and cynomolgus macaque monkeys following five-hour treatment with phorbol myristic acetate (PMA) and Ca++ ionophore (A23187) in the presence of monensin. The staining pattern of 4S.B3 in CD3+ cells is similar to that observed with peripheral blood T lymphocytes from normal human donors. This reagent is useful for intracellular immunofluorescent staining for flow cytometric analysis to identify and enumerate IFN-γ + cells within a mixed cell population.
- Format PE
- Excitation Source Blue 488 nm,Green 532 nm,Yellow/Green 561 nm
- Excitation Max 496 nm
- Emission Max 578 nm
R-phycoerythrin (PE) is an accessory photosynthetic pigment found in red algae. It exists in vitro as a 240-kDa protein with 23 phycoerythrobilin chromophores per molecule. This makes PE one of the brightest fluorochromes for flow cytometry applications, but its photobleaching properties make it unsuitable for fluorescence microscopy.
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Preparation and Storage
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.
The antibody was conjugated with R-PE under optimum conditions, and unconjugated antibody and free PE were removed.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- An isotype control should be used at the same concentration as the antibody of interest.
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Species testing during development may have been performed with a different format of the same clone. Selected applications have been tested for cross-reactivity.
- Please refer to www.bdbiosciences.com/pharmingen/protocols for technical protocols.
Immunofluorescent Staining for Flow Cytometric Analysis: The PE Mouse Anti-Human IFN-γ antibody is useful for immunofluorescent staining and flow cytometric analysis to identify and enumerate IFN-γ producing cells within mixed cell populations. For optimal immunofluorescent staining for flow cytometric analysis, the anti-cytokine antibody should be titrated (≤ 0.5 µg mAb/million cells). For specific methodology, please visit our web site, http://www.bdbiosciences.com/us/s/resources, and go to the protocols section under "Intracellular Flow".
A useful control for demonstrating specificity of staining is either of the following: 1) pre-block the PE Mouse Anti-Human IFN-γ antibody with ligand (e.g., rhIFN-γ, Cat. No. 554617) prior to staining, or 2) pre-block the fixed/permeabilized cells with Purified Mouse Anti-Human IFN-γ (Cat. No. 554549) prior to staining. The staining technique and blocking controls are described in detail by C. Prussin and D. Metcalfe. A suitable mouse IgG1 isotype control for assessing the level of background staining on paraformaldehyde-fixed/saponin-permeabilized human cells is PE Mouse IgG1, κ Isotype Control (Cat. No. 554680); use at comparable concentrations to antibody of interest (e.g., ≤ 0.5 µg mAb/1 million cells).