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BV421 Mouse Anti-Human TNF
BV421 Mouse Anti-Human TNF
Multiparameter flow cytometric analysis of TNF expressed in stimulated human peripheral blood mononuclear cells. HiCK-1 Human Cytokine Positive Control Cells (Cat. No. 555061) were permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained with either BD Horizon™ BV421 Mouse IgG1, κ Isotype Control  (Cat. No. 562438; Left Panel) or BD Horizon™ BV421 Mouse Anti-Human TNF antibody (Cat. No. 562783/566275; Middle Panel). To demonstrate specificity of staining, the fixed and permeabilized cells were preincubated with Purified Mouse Anti-Human TNF antibody (10 µg, Cat. No. 554510; Right Panel) to block subsequent staining with the BV421 Mouse Anti-Human TNF antibody. Two-color flow cytometric dot plots show the correlated expression patterns of TNF, Ig Isotype control or blocked TNF staining versus autofluorescence for gated events with the forward and side light-scatter characteristics of intact peripheral blood mononuclear cells. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Multiparameter flow cytometric analysis of TNF expressed in stimulated human peripheral blood mononuclear cells. HiCK-1 Human Cytokine Positive Control Cells (Cat. No. 555061) were permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained with either BD Horizon™ BV421 Mouse IgG1, κ Isotype Control  (Cat. No. 562438; Left Panel) or BD Horizon™ BV421 Mouse Anti-Human TNF antibody (Cat. No. 562783/566275; Middle Panel). To demonstrate specificity of staining, the fixed and permeabilized cells were preincubated with Purified Mouse Anti-Human TNF antibody (10 µg, Cat. No. 554510; Right Panel) to block subsequent staining with the BV421 Mouse Anti-Human TNF antibody. Two-color flow cytometric dot plots show the correlated expression patterns of TNF, Ig Isotype control or blocked TNF staining versus autofluorescence for gated events with the forward and side light-scatter characteristics of intact peripheral blood mononuclear cells. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Product Details
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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
AB_2737790
Aqueous buffered solution containing BSA and ≤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 with BD Horizon BV421 under optimum conditions, and unconjugated antibody and free BD Horizon BV421 were removed.

Recommended Assay Procedures

For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes are used in the same experiment.  Fluorescent dye interactions may cause staining artifacts which may affect data interpretation.  The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions.  More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349).

Product Notices

  1. 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).
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  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. Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
  7. BD Horizon Brilliant Violet 421 is covered by one or more of the following US patents: 8,158,444; 8,362,193; 8,575,303; 8,354,239.
  8. BD Horizon Brilliant Stain Buffer is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,575,303; 8,354,239.
  9. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  10. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
566275 Rev. 2
Antibody Details
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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.

The antibody was conjugated to BD Horizon™ BV421 which is part of the BD Horizon Brilliant™ Violet family of dyes. With an Ex Max of 407-nm and Em Max at 421-nm, BD Horizon™ BV421 can be excited by the violet laser and detected in the standard Pacific Blue™ filter set (eg, 450/50-nm filter). BD Horizon™ BV421 conjugates are very bright, often exhibiting a 10 fold improvement in brightness compared to Pacific Blue™ conjugates.

566275 Rev. 2
Format Details
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BV421
The BD Horizon Brilliant Violet™ 421 (BV421) Dye is part of the BD Horizon Brilliant Violet™ family of dyes. This polymer-technology based dye has an excitation maximum (Ex Max) of 407-nm and an emission maximum (Em Max) at 423-nm. Driven by BD innovation, BV421 is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 420-nm (e.g., a 431/28-nm or 450/50-nm bandpass filter). BV421 is an ideal alternative for V450 as it is approximately ten times brighter with less spillover into the BV510/V500 detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BV421
Violet 405 nm
407 nm
423 nm
566275 Rev.2
Citations & References
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Development References (13)

  1. Black RA, Rauch CT, Kozlosky CJ, et al. A metalloproteinase disintegrin that releases tumour-necrosis factor-alpha from cells. Nature. 1997; 385(6618):729-733. (Biology). View Reference
  2. 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
  3. Jaattela, M. . Biologic activities and mechanisms of action of tumor necrosis factor-α/cachectin. Lab Invest. 1991; 64:724-742. (Biology).
  4. 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. (Biology). View Reference
  5. Kriegler M, Perez C, DeFay K, Albert I, Lu SD. A novel form of TNF/cachectin is a cell surface cytotoxic transmembrane protein: ramifications for the complex physiology of TNF. Cell. 1988; 53(1):45-53. (Biology). View Reference
  6. Petyovka N, Lyach L, Voitenok NN. Homologous ELISA for detection of oligomeric human TNF: properties of the assay. J Immunol Methods. 1995; 186(2):161-170. (Biology). View Reference
  7. 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. (Methodology: IC/FCM Block). View Reference
  8. 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
  9. 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
  10. Smith RA, Baglioni C. The active form of tumor necrosis factor is a trimer. J Biol Chem. 1987; 262(15):6951-6954. (Biology). View Reference
  11. Sopper S, Stahl-Hennig C, Demuth M, Johnston IC, Dorries R, ter Meulen V. Lymphocyte subsets and expression of differentiation markers in blood and lymphoid organs of rhesus monkeys. Cytometry. 1997; 29(4):351-362. (Biology). View Reference
  12. Verdier F, Aujoulat M, Condevaux F, Descotes J. Determination of lymphocyte subsets and cytokine levels in cynomolgus monkeys. Toxicology. 1995; 105(1):81-90. (Biology). View Reference
  13. Wang AM, Creasey AA, Ladner MB, et al. Molecular cloning of the complementary DNA for human tumor necrosis factor. Science. 1985; 228(4696):149-154. (Biology). View Reference
View All (13) View Less
566275 Rev. 2

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


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