Skip to main content Skip to navigation
Hamburger Menu
Search icon
Country Selector Country Selector
Luxembourg (English)
Profile Icon Profile Icon
Sign-in/Register UserName
Products

Solutions

Discover & Learn

Resources & Tools

Support

Search icon Search icon
Close Menu
      Alert Icon
      RY586 Mouse Anti-Human TNF
      RY586 Mouse Anti-Human TNF
      Flow cytometric analysis of TNF expression in stimulated Human peripheral blood lymphocytes. Human peripheral blood mononuclear cells (PBMC) were stimulated for 5 hours with Phorbol 12-Myristate 13-Acetate (Sigma P-8139; 50 ng/ml final concentration) and Ionomycin (Sigma I-0634; 1 μg/ml final concentration) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) (Cat. No. 554724). The cells were harvested, washed with BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656) and fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) followed by washing and permeabilization using BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained in BD Perm/Wash™ Buffer with BD Horizon™ BV421 Mouse Anti-Human CD3 antibody (Cat. No. 562426) and with either BD Horizon™ RY586 Mouse IgG1, κ Isotype Control (Cat No. 568097, Left Plot) or with BD Horizon™ RY586 Mouse Anti-Human TNF antibody (Cat No. 568461/568462, Right Plot). The pseudocolor density plot showing the correlated expression of TNF (or Ig Isotype control staining) versus CD3 was derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Flow Cytometer System and FlowJo™ software.
      RY586 Mouse Anti-Human TNF
      Flow cytometric analysis of TNF expression in stimulated Human peripheral blood lymphocytes. Human peripheral blood mononuclear cells (PBMC) were stimulated for 5 hours with Phorbol 12-Myristate 13-Acetate (Sigma P-8139; 50 ng/ml final concentration) and Ionomycin (Sigma I-0634; 1 μg/ml final concentration) in the presence of BD GolgiStop™ Protein Transport Inhibitor (containing Monensin) (Cat. No. 554724). The cells were harvested, washed with BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656) and fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) followed by washing and permeabilization using BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained in BD Perm/Wash™ Buffer with BD Horizon™ BV421 Mouse Anti-Human CD3 antibody (Cat. No. 562426) and with either BD Horizon™ RY586 Mouse IgG1, κ Isotype Control (Cat No. 568097, Left Plot) or with BD Horizon™ RY586 Mouse Anti-Human TNF antibody (Cat No. 568461/568462, Right Plot). The pseudocolor density plot showing the correlated expression of TNF (or Ig Isotype control staining) versus CD3 was derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Flow Cytometer System and FlowJo™ software.
      Product Details
      Down Arrow Up Arrow


      BD Horizon™
      Tumor necrosis factor alpha; TNF-a; TNF-α; TNFSF2; Cachectin
      Human (QC Testing), Rhesus, Cynomolgus, Baboon (Tested in Development)
      Mouse IgG1, κ
      Recombinant Human TNF
      Intracellular staining (flow cytometry) (Routinely Tested)
      5 µl
      Aqueous buffered solution containing ≤0.09% sodium azide.
      RUO


      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 to the dye under optimum conditions and unreacted dye was removed.
      Show More blue down arrow Show Less blue up arrow

      Recommended Assay Procedures

      BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.

      Show More blue down arrow Show Less blue up arrow

      Product Notices

      1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      2. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
      3. An isotype control should be used at the same concentration as the antibody of interest.
      4. 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.
      5. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      6. CF™ is a trademark of Biotium, Inc.
      7. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
      8. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
      9. Human donor specific background has been observed in relation to the presence of anti-polyethylene glycol (PEG) antibodies, developed as a result of certain vaccines containing PEG, including some COVID-19 vaccines. We recommend use of BD Horizon Brilliant™ Stain Buffer in your experiments to help mitigate potential background. For more information visit https://www.bdbiosciences.com/en-us/support/product-notices.
      Show More blue down arrow Show Less blue up arrow
      Antibody Details
      Down Arrow Up Arrow
      MAb11

      The MAb11 monoclonal antibody specifically binds to human tumor necrosis factor (TNF, also known as TNF-α) protein. TNF is an efficient juxtacrine, paracrine and endocrine mediator of inflammatory and immune functions. It regulates the growth and differentiation of a variety of cell types. TNF is cytotoxic for transformed cells when in conjunction with IFN-γ. It is secreted by activated monocytes/macrophages and other cells such as B cells, T cells and fibroblasts. The immunogen used to generate the MAb11 hybridoma was recombinant human TNF. The MAb11 antibody has been reported to crossreact with Rhesus Macaque TNF.

      Format Details
      Down Arrow Up Arrow
      RY586
      The BD Horizon RealYellow™ 586 (RY586) Dye is part of the BD family of yellow-green dyes. It is a small organic fluorochrome with an excitation maximum (Ex Max) at 565-nm and an emission maximum (Em Max) at 586-nm. Driven by BD innovation, RY586 can be used on both spectral and conventional cytometers and is designed to be excited by the Yellow-Green laser (561-nm) with minimal excitation by the 488-nm Blue laser. For conventional instruments equipped with a Yellow-Green laser (561-nm), RY586 can be used as an alternative to PE and we recommend using an optical filter centered near 586-nm (eg, a 586/15-nm bandpass filter). For spectral instruments equipped with a Yellow-Green laser (561-nm), it can be used in conjunction with PE. Compared to PE, RY586 is similar in brightness, minimal spillover into Blue detectors, and increased spillover into the 610/20-nm (PE-CF594) detector. Please ensure that your instrument configuration (lasers and optical filters) is appropriate for this dye.
      altImg
      RY586
      Yellow-Green 561 nm
      564 nm
      586 nm
      Citations & References
      Down Arrow Up Arrow
      View product citations for antibody "568462" on CiteAb

      Development References (12)

      1. Andersen H, Rossio JL, Coalter V, et al. Characterization of rhesus macaque natural killer activity against a rhesus-derived target cell line at the single-cell level.. Cell Immunol. 231(1-2):85-95. (Clone-specific: Flow cytometry). View Reference
      2. Castro I, Giret TM, Magnani DM, et al. Cellular Immune Responses against Simian T-Lymphotropic Virus Type 1 Target Tax in Infected Baboons.. J Virol. 2016; 90(11):5280-5291. (Clone-specific: Flow cytometry). View Reference
      3. Danis VA, Franic GM, Rathjen DA, Brooks PM. Effects of granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-2, interferon-gamma (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha) and IL-6 on the production of immunoreactive IL-1 and TNF-alpha by human monocytes. Clin Exp Immunol. 1991; 85(1):143-150. (Clone-specific: ELISA). View Reference
      4. Engele M, Stössel E, Castiglione K, et al. Induction of TNF in human alveolar macrophages as a potential evasion mechanism of virulent Mycobacterium tuberculosis.. J Immunol. 2002; 168(3):1328-37. (Clone-specific: Flow cytometry). View Reference
      5. Foulds KE, Donaldson M, Roederer M. OMIP-005: Quality and phenotype of antigen-responsive rhesus macaque T cells. Cytometry A. 2012; 81(5):360-361. (Clone-specific: Flow cytometry). View Reference
      6. Geisbert TW, Bailey M, Geisbert JB, et al. Vector choice determines immunogenicity and potency of genetic vaccines against Angola Marburg virus in nonhuman primates.. J Virol. 2010; 84(19):10386-94. (Clone-specific: Flow cytometry). View Reference
      7. Jason J, Larned J. Single-cell cytokine profiles in normal humans: comparison of flow cytometric reagents and stimulation protocols. J Immunol Methods. 1997; 207(1):13-22. (Clone-specific: Flow cytometry). View Reference
      8. Lecoeur H, Ledru E, Gougeon ML. A cytofluorometric method for the simultaneous detection of both intracellular and surface antigens of apoptotic peripheral lymphocytes.. J Immunol Methods. 1998; 217(1-2):11-26. (Clone-specific: Flow cytometry). View Reference
      9. Mascher B, Schlenke P, Seyfarth M. Expression and kinetics of cytokines determined by intracellular staining using flow cytometry. J Immunol Methods. 1999; 223(1):115-121. (Clone-specific: Flow cytometry). View Reference
      10. Moris P, Bellanger A, Ofori-Anyinam O, Jongert E, Yarzabal Rodriguez JP, Janssens M. Whole blood can be used as an alternative to isolated peripheral blood mononuclear cells to measure in vitro specific T-cell responses in human samples.. J Immunol Methods. 2021; 492:112940. (Clone-specific: Flow cytometry). View Reference
      11. Rathjen DA, Cowan K, Furphy LJ, Aston R. Antigenic structure of human tumour necrosis factor: recognition of distinct regions of TNF alpha by different tumour cell receptors. Mol Immunol. 1991; 28(1-2):79-86. (Clone-specific: ELISA). View Reference
      12. Sander B, Andersson J, Andersson U. Assessment of cytokines by immunofluorescence and the paraformaldehyde-saponin procedure. Immunol Rev. 1991; 119:65-93. (Biology). View Reference
      View All (12) View Less

      Please refer to Support Documents for Quality Certificates


      Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described


      Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

      For Research Use Only. Not for use in diagnostic or therapeutic procedures.