Skip to main content Skip to navigation
V450 Rat Anti-Mouse TNF
V450 Rat Anti-Mouse TNF

Flow cytometric analysis for TNF in activated mouse splenocytes.  Mouse Intracellular Cytokine-1 positive control cells (MiCK-1) offered by BD Biosciences as MN 554652, are activated mouse splenocytes prepared in the presence of a protein transport inhibitor.  Fixed and permeabilized MiCK-1 cells were stained either with a BD Horizon™ V450 Rat IgG1, κ isotype control (left panel) or with the BD Horizon™ Rat Anti-Mouse TNF antibody (right panel).  Dot plots were derived from gated events based on light scattering characteristics for lymphocytes.  Flow cytometry was performed on a BD™ LSR II flow cytometry system.

Flow cytometric analysis for TNF in activated mouse splenocytes.  Mouse Intracellular Cytokine-1 positive control cells (MiCK-1) offered by BD Biosciences as MN 554652, are activated mouse splenocytes prepared in the presence of a protein transport inhibitor.  Fixed and permeabilized MiCK-1 cells were stained either with a BD Horizon™ V450 Rat IgG1, κ isotype control (left panel) or with the BD Horizon™ Rat Anti-Mouse TNF antibody (right panel).  Dot plots were derived from gated events based on light scattering characteristics for lymphocytes.  Flow cytometry was performed on a BD™ LSR II flow cytometry system.

Product Details
Down Arrow Up Arrow


BD Horizon™
Mouse (QC Testing)
Rat IgG1
Recombinant Mouse TNF
Intracellular staining (flow cytometry) (Routinely Tested)
0.2 mg/ml
AB_1727581
Aqueous buffered solution containing protein stabilizer 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™ V450 under optimum conditions, and unreacted BD Horizon™ V450 was removed.

Recommended Assay Procedures

Flow cytometry:  The MP6-XT22 antibody is useful for immunofluorescent staining and flow cytometric analysis to identify and enumerate TNF producing cells within mixed cell populations.  A useful control investigators may consider using for demonstrating specificity of staining, is to pre-block with one of the following reagents: (1) recombinant mouse TNF (Cat. No. 554589) or (2) unlabeled MP6-XT22 antibody (Cat. No. 554416), prior to staining.

Cell Preparation: Investigators not wishing to utilize MiCK-1 cells may alternatively prepare mouse splenocytes (e.g BALB/c) stimulated for 4-6 hours with PMA (5 ng/mL, Sigma-Aldrich Cat. No. P-8139) and ionomycin (500 ng, Sigma-Aldrich Cat. No. I-0634) in the presence of 1 µg/mL Brefeldin A (BD GolgiPlug™ MN 555029).  Investigators are advised to fix and permeabilize the cells prior to staining.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. BD Horizon V450 has a maximum absorption of 406 nm and maximum emission of 450 nm. Before staining with this reagent, please confirm that your flow cytometer is capable of exciting the fluorochrome and discriminating the resulting fluorescence.
  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. Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
  6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  7. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
560655 Rev. 1
Antibody Details
Down Arrow Up Arrow
MP6-XT22

The MP6-XT22 antibody specifically binds to mouse Tumor Necrosis Factor (TNF, also known as TNF-α).  TNF is produced by many activated cell types including monocytes, macrophages, astrocytes, granulocytes, mast cells, T and B lymphocytes, NK cells, keratinocytes, fibroblasts, adipocytes, and certain tumor cells. Activated cells express type II transmembrane TNF glycoproteins that associate as homotrimeric complexes. After enzymatic cleavage, the extracellular regions of membrane TNF are shed as soluble homotrimers. TNF is a potent multifunctional cytokine that can exert regulatory and cytotoxic effects on a wide range of normal lymphoid and non-lymphoid cells and tumor cells. Although TNF serves as a primary mediator in protective immune responses against microbial and viral pathogens, it can also drive systemic pathophysiologic responses including septic shock, cachexia and autoimmune diseases. Mouse TNF exerts its biological activities by binding and signaling through cell surface membrane Type I and Type II TNF Receptors (aka, TNFRI/CD120a and TNFRII/CD120b, respectively).

The antibody is conjugated to BD Horizon™ V450, which has been developed for use in multicolor flow cytometry experiments and is available exclusively from BD Biosciences. It is excited by the Violet laser Ex max of 406 nm and has an Em Max at 450 nm. Conjugates with BD Horizon™ V450 can be used in place of Pacific Blue™ conjugates.

560655 Rev. 1
Format Details
Down Arrow Up Arrow
V450
BD Horizon™ V450 Dye is part of the BD Horizon™ violet family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 405-nm and an emission maximum (Em Max) at 450-nm. BD Horizon™ V450, driven by BD innovation, is designed to be excited by the violet laser (405 nm) and detected using an optical filter centered near 450-nm (e.g., a 450/50-nm bandpass filter). The dye can be excited by the UV (355-nm) laser resulting in cross-laser excitation and spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
altImg
V450
Violet 405 nm
405 nm
450 nm
560655 Rev.1
Citations & References
Down Arrow Up Arrow

Development References (13)

  1. Abrams J. Immunoenzymetric assay of mouse and human cytokines using NIP-labeled anti-cytokine antibodies. Curr Protoc Immunol. 2001; 1:6.20-6.21. (Biology). View Reference
  2. Abrams JS, Roncarolo MG, Yssel H, Andersson U, Gleich GJ, Silver JE. Strategies of anti-cytokine monoclonal antibody development: immunoassay of IL-10 and IL-5 in clinical samples. Immunol Rev. 1992; 127:5-24. (Biology). View Reference
  3. Drew PD, Chavis JA. Female sex steroids: effects upon microglial cell activation. J Neuroimmunol. 2000; 111(1-2):77-85. (Biology). View Reference
  4. Drew PD, Chavis JA. Inhibition of microglial cell activation by cortisol. Brain Res Bull. 2000; 52(5):391-396. (Biology). View Reference
  5. Drew PD, Chavis JA.. The cyclopentone prostaglandin 15-deoxy-Delta(12,14) prostaglandin J2 represses nitric oxide, TNF-alpha, and IL-12 production by microglial cells.. J Neuroimmunol. 2001; 115(1-2):28-35. (Biology). View Reference
  6. Ferran C, Dautry F, Merite S, et al. Anti-tumor necrosis factor modulates anti-CD3-triggered T cell cytokine gene expression in vivo. J Clin Invest. 1994; 93(5):2189-2196. (Biology). View Reference
  7. He J, Gurunathan S, Iwasaki A, Ash-Shaheed B, Kelsall BL. Primary role for Gi protein signaling in the regulation of interleukin 12 production and the induction of T helper cell type 1 responses . J Exp Med. 2000; 191(9):1605-1610. (Biology). View Reference
  8. Leiby DA, Fortier AH, Crawford RM, Schreiber RD, Nacy CA. In vivo modulation of the murine immune response to Francisella tularensis LVS by administration of anticytokine antibodies. Infect Immun. 1992; 60(1):84-89. (Biology). View Reference
  9. Merrick BA, He CY, Craig WA, et al. Two dimensional gel electrophoresis of cellular and secreted proteins from rat alveolar macrophages after lipopolysaccharide treatment. Appl Theor Electrophor. 1992; 2(6):177-187. (Biology). View Reference
  10. 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: Flow cytometry). View Reference
  11. Rabinovici R, Bugelski PJ, Esser KM, et al. Tumor necrosis factor-alpha mediates endotoxin-induced lung injury in platelet activating factor-primed rats. J Pharmacol Exp Ther. 1993; 267(3):1550-1557. (Biology). View Reference
  12. Sheehan KC, Ruddle NH, Schreiber RD. Generation and characterization of hamster monoclonal antibodies that neutralize murine tumor necrosis factors. J Immunol. 1989; 142(11):3884-3893. (Biology). View Reference
  13. Takahashi S, Kapas L, Fang J, Krueger JM. An anti-tumor necrosis factor antibody suppresses sleep in rats and rabbits. Brain Res. 1995; 690(2):241-244. (Biology). View Reference
View All (13) View Less
560655 Rev. 1

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.