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PE Mouse Anti-Human GITR (CD357)
PE Mouse Anti-Human GITR (CD357)

Multiparameter flow cytometric analysis of GITR (CD357) expression on human peripheral blood lymphocytes. PBMC were cultured for 3 days in medium without (Unstimulated; Left Plot) or with Phytohemagglutinin-M (PHA-M Stimulated; 1.5% v/v; Right Plot). Cells were stained with BD Horizon™ BUV395 Mouse Anti-Human CD4 antibody (Cat. No. 564724) and with either PE Mouse IgG2b, κ Isotype Control (Cat. No. 555058; no staining, data not shown) or PE Mouse Anti-Human GITR (CD357) antibody (Cat. No. 567191/567192). BD Via-Probe™ Cell Viability 7-AAD Solution (Cat. No. 555815/555816) was added to cells right before analysis. The bivariate pseudocolor density plots showing the correlated expression of GITR (CD357) versus CD4 were derived from gated events with the forward and side light-scatter characteristics of viable (7-AAD) human peripheral blood lymphocytes.

Multiparameter flow cytometric analysis of GITR (CD357) expression on human CD4+ peripheral blood lymphocytes. PBMC were stained with BD Horizon™ BUV395 Mouse Anti-Human CD4 (Cat. No. 564724) and APC Mouse Anti-Human CD25 (Cat. No. 555434) antibodies and with either PE Mouse IgG2b, κ Isotype Control (Cat. No. 555058; dashed line histograms) or PE Mouse Anti-Human GITR (CD357) antibody (Cat. No. 567191/567192; solid line histograms). BD Via-Probe™ Cell Viability 7-AAD Solution was added to cells right before analysis. Histograms showing the expression of GITR (CD357) [or Ig Isotype control staining] were derived from CD4+CD25-negative (Left Plot) or CD4+CD25-bright positive (Right Plot) gated events with the forward and side light-scatter characteristics of viable (7-AAD negative) lymphocytes.

   Flow cytometry and data analysis were performed using a BD LSRFortessa™ Cell Analyzer System and FlowJo™ software.

Multiparameter flow cytometric analysis of GITR (CD357) expression on human peripheral blood lymphocytes. PBMC were cultured for 3 days in medium without (Unstimulated; Left Plot) or with Phytohemagglutinin-M (PHA-M Stimulated; 1.5% v/v; Right Plot). Cells were stained with BD Horizon™ BUV395 Mouse Anti-Human CD4 antibody (Cat. No. 564724) and with either PE Mouse IgG2b, κ Isotype Control (Cat. No. 555058; no staining, data not shown) or PE Mouse Anti-Human GITR (CD357) antibody (Cat. No. 567191/567192). BD Via-Probe™ Cell Viability 7-AAD Solution (Cat. No. 555815/555816) was added to cells right before analysis. The bivariate pseudocolor density plots showing the correlated expression of GITR (CD357) versus CD4 were derived from gated events with the forward and side light-scatter characteristics of viable (7-AAD) human peripheral blood lymphocytes.

Multiparameter flow cytometric analysis of GITR (CD357) expression on human CD4+ peripheral blood lymphocytes. PBMC were stained with BD Horizon™ BUV395 Mouse Anti-Human CD4 (Cat. No. 564724) and APC Mouse Anti-Human CD25 (Cat. No. 555434) antibodies and with either PE Mouse IgG2b, κ Isotype Control (Cat. No. 555058; dashed line histograms) or PE Mouse Anti-Human GITR (CD357) antibody (Cat. No. 567191/567192; solid line histograms). BD Via-Probe™ Cell Viability 7-AAD Solution was added to cells right before analysis. Histograms showing the expression of GITR (CD357) [or Ig Isotype control staining] were derived from CD4+CD25-negative (Left Plot) or CD4+CD25-bright positive (Right Plot) gated events with the forward and side light-scatter characteristics of viable (7-AAD negative) lymphocytes.

   Flow cytometry and data analysis were performed using a BD LSRFortessa™ Cell Analyzer System and FlowJo™ software.

Product Details
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BD Pharmingen™
GITR-D; TNFRSF18; activation-inducible TNFR family receptor; AITR
Human (QC Testing)
Mouse BALB/c IgG2b, κ
Human GITR Recombinant Protein
Flow cytometry (Routinely Tested)
5 µl
8784
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 to the dye under optimum conditions and unconjugated antibody and free dye were removed.

Recommended Assay Procedures

BD™ CompBeads can be used as surrogates to assess fluorescence spillover (Compensation).  When fluorochrome conjugated antibodies are bound to 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 cell and CompBead to ensure that BD Comp beads are appropriate for your specific cellular application.

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. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  7. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
567192 Rev. 1
Antibody Details
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V27-580

The V27-580 monoclonal antibody specifically binds to GITR (Glucocorticoid-Induced Tumor necrosis factor Receptor), a member of the tumor necrosis factor receptor (TNFR) superfamily that is designated TNFRSF18. In the human, GITR is expressed at low levels in peripheral blood T cells, bone marrow, thymus, spleen, and lymph nodes and is up-regulated upon antigen stimulation or by treatment with anti-CD3 plus anti-CD28. GITR is also reported to be constitutively expressed on Treg cells. GITR's ligand (GITRL) is a member of the TNF superfamily, is designated TNFSF18, and is expressed on antigen presenting cells. The GITR cytoplasmic domain has striking homology with the cytoplasmic domains of the co-stimulatory receptors CD137 (4-1BB), CD134 (OX40) and CD27. GITR signaling is mediated by signaling adaptors, TNFR-associated factors (TRAFs), that affect signaling pathways (eg, Erk, JNK, MAPK and NF-κB) to enhance T-cell survival and cytokine production. The effects of GITR signaling upon the dynamic and interconnected roles of effector and regulatory T lymphocytes in the immune response are under investigation.

                

567192 Rev. 1
Format Details
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PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
altImg
PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
567192 Rev.1
Citations & References
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Development References (7)

  1. Clouthier DL, Watts TH. Cell-specific and context-dependent effects of GITR in cancer, autoimmunity, and infection.. Cytokine Growth Factor Rev. 2014; 25(2):91-106. (Biology). View Reference
  2. Kwon B, Yu KY, Ni J, et al. Identification of a novel activation-inducible protein of the tumor necrosis factor receptor superfamily and its ligand.. J Biol Chem. 1999; 274(10):6056-61. (Biology). View Reference
  3. Li Z, Mahesh SP, Kim BJ, Buggage RR, Nussenblatt RB. Expression of glucocorticoid induced TNF receptor family related protein (GITR) on peripheral T cells from normal human donors and patients with non-infectious uveitis.. J Autoimmun. 2003; 21(1):83-92. (Biology). View Reference
  4. Nocentini G, Giunchi L, Ronchetti S, et al. A new member of the tumor necrosis factor / nerve growth factor receptor family inhibits T cell receptor-induced apoptosis... Proc Natl Acad Sci U S A. 1997; 94(12):6216-6221. (Biology). View Reference
  5. Pedroza-Gonzalez A, Zhou G, Singh SP, et al. GITR engagement in combination with CTLA-4 blockade completely abrogates immunosuppression mediated by human liver tumor-derived regulatory T cells ex vivo.. Oncoimmunology. 2015; 4(12):e1051297. (Biology). View Reference
  6. Placke T, Kopp HG, Salih HR. Glucocorticoid-induced TNFR-related (GITR) protein and its ligand in antitumor immunity: functional role and therapeutic modulation.. Clin Dev Immunol. 2010; 2010:239083. (Biology). View Reference
  7. Shevach EM, Stephens GL. The GITR-GITRL interaction: co-stimulation or contrasuppression of regulatory activity?. Nat Rev Immunol. 2006; 6(8):613-8. (Biology). View Reference
View All (7) View Less
567192 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.

Refer to manufacturer's instructions for use and related User Manuals and Technical Data Sheets before using this product 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.