Preparation And Storage
Recommended Assay Procedures
ELISA Detection: Biotin Rat Anti-Human CD120b (Cat. No. 552477) serves as the detection antibody in a sandwich ELISA for measuring human TNFRII protein levels. Biotin Rat Anti-Human CD120b (Cat. No. 552477) can be paired with purified MR2-2 anti-human TNFRII using the recombinant human soluble TNFRII as the standard. This detection antibody should be titrated between 0.5 -1 µg/ml to determine its optimal concentration for ELISA detection. To obtain linear standard curves, doubling dilutions of recombinant soluble human TNFRII ranging from 1000 to 10 pg/ml are recommended for inclusion in each ELISA plate. For specific methodology, please visit our website, http://www.bdbiosciences.com/us/s/resources, and go to the protocols section under "ELISA and ELISPOT".
Note: This ELISA antibody pair shows no cross-reactivity with the following recombinant human cytokines: CD14,CD40, CD40L, IL-1Ra, IL-1RII, IL-1α, IL-1β, IL-2, IL-3, IL-4, sIL-4R, IL-5, IL-6, IL-6R, IL-7, IL-8, IL-10, IL-12p40, IL-12p70, IL-13, IL-15, TNF, LT-α, sTNFRI, IFNγ, TRAIL, GM-CSF, TGFβ, Trx. This ELISA antibody pair also shows no cross-reactivity with recombinant mouse sTNFRI, recombinant mouse sTNFRII, recombinant mouse TNF.
Immunofluorescent Staining and Flow Cytometric Analysis: The Purified Rat Anti-Human CD120b (Cat. No. 551311) antibody can be used for the immunofluorescent staining (≤ 1 µg antibody/10e6 cells) and flow cytometric analysis of human nucleated cells to measure their expressed levels of surface TNFRII. An appropriate immunoglobulin is Purified Rat IgG2b, κ Isotype Control (Cat. No. 555846). Other formats available for immunofluorescent staining include, PE Rat Anti-Human CD120b (Cat. No. 552418). Please note also that as a consequence of in vivo or in vitro activation, cell surface TNFRII can either be shed by cells or transiently expressed at higher levels. As a result, cellular activation can affect the cell's overall expressed level of surface TNFRII.
Product Notices
- 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.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
The hTNFR-M1 antibody specifically recognizes the extracellular domain of the 75 kDa transmembrane receptor for the human cytokines, tumor necrosis factor (TNF or TNF-α) and lymphotoxin-alpha (LT-α3, aka, lymphotoxin or TNF-β). This receptor is referred to as the p75 or Type II Tumor Necrosis Factor Receptor (TNFRII) [aka, CD120b]. Human TNFRII proteins are expressed by hematopoietic cells including macrophages, neutrophils, lymphocytes, thymocytes and mast cells. TNFRII is expressed by a variety of other cell types including endothelial cells, cardiac myocytes and prostate cells. Naive B cells express very low or undetectable levels of TNFRII whereas mature erythrocytes and platelets are uniformly negative for TNFRII expression. The immunogen used to generate the hTNFR-M1 hybridoma was COS- expressed recombinant human TNFRII.
Development References (10)
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Erikstein BK, Smeland EB, Blomhoff HK, et al. Independent regulation of 55-kDa and 75-kDa tumor necrosis factor receptors during activation of human peripheral blood B lymphocytes. Eur J Immunol. 1991; 21(4):1033-1037. (Biology). View Reference
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Hohmann HP, Remy R, Brockhaus M, van Loon AP. Two different cell types have different major receptors for human tumor necrosis factor (TNF alpha). J Biol Chem. 1989; 264(25):14927-14934. (Biology). View Reference
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Munker R, DiPersio J, Koeffler HP. Tumor necrosis factor: receptors on hematopoietic cells. Blood. 1987; 70(6):1730-1734. (Clone-specific: Flow cytometry). View Reference
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Wallach D, Engelmann H, Nophar Y, et al. Soluble and cell surface receptors for tumor necrosis factor. Agents Actions Suppl. 1991; 35:51-57. (Biology). View Reference
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Ware CF, Crowe PD, Vanarsdale TL, et al. Tumor necrosis factor (TNF) receptor expression in T lymphocytes. Differential regulation of the type I TNF receptor during activation of resting and effector T cells.. J Immunol. 1991; 147(12):4229-38. (Biology). View Reference
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Zola H. Detection of cytokine receptors by flow cytometry. In: Coligan JE, Kruisbeek AM, Margulies DH, Shevach EM, Strober W, ed. Current Protocols in Immunology. New York: Green Publishing Associates and Wiley-Interscience; 1995:6.21.1-6.21.18.
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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
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.
