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      RB613 Mouse Anti-Stat1 (pY701)
      RB613 Mouse Anti-Stat1 (pY701)
      Flow cytometric analysis of Stat1 (pY701) expression in Unstimulated and Stimulated Human peripheral blood monocytes.  Human peripheral blood mononuclear cells (PBMC) were either left Unstimulated (dashed line histogram) or Stimulated (solid line histogram) with 100 ng/ml (final concentration) of BD Pharmingen™ Recombinant Human IFN-γ protein (Cat. No. 554617) for 15 minutes at 37°C. The PBMC were fixed using BD Cytofix™ Fixation buffer (Cat. No. 554655) for 10 minutes at 37°C, then permeabilized using BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice for 30 minutes. The cells were washed twice with stain buffer and then stained with BD Phosflow™ RB613 Mouse Anti-Stat1 (pY701) antibody (Cat. No. 571245/571309). Flow cytometry and data analysis were performed using a 5-laser Flow Cytometer System and FlowJo™ Software.
      RB613 Mouse Anti-Stat1 (pY701)
      Flow cytometric analysis of Stat1 (pY701) expression in Unstimulated and Stimulated Human peripheral blood monocytes.  Human peripheral blood mononuclear cells (PBMC) were either left Unstimulated (dashed line histogram) or Stimulated (solid line histogram) with 100 ng/ml (final concentration) of BD Pharmingen™ Recombinant Human IFN-γ protein (Cat. No. 554617) for 15 minutes at 37°C. The PBMC were fixed using BD Cytofix™ Fixation buffer (Cat. No. 554655) for 10 minutes at 37°C, then permeabilized using BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice for 30 minutes. The cells were washed twice with stain buffer and then stained with BD Phosflow™ RB613 Mouse Anti-Stat1 (pY701) antibody (Cat. No. 571245/571309). Flow cytometry and data analysis were performed using a 5-laser Flow Cytometer System and FlowJo™ Software.
      Product Details
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      BD Phosflow™
      Signal Transducer and Activator of Transcription-1; STAT91; ISGF-3; CANDF7
      Human (QC Testing), Mouse (Tested in Development)
      Mouse IgG2a
      Phosphorylated Human Stat1 Peptide
      Intracellular staining (flow cytometry) (Routinely Tested)
      5 µl/test
      Aqueous buffered solution containing ≤0.09% sodium azide.
      RUO


      Preparation And Storage

      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. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
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      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.

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      Product Notices

      1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      2. 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.
      3. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
      4. 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).
      5. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      6. An isotype control should be used at the same concentration as the antibody of interest.
      7. This product is provided under an intellectual property license between Life Technologies Corporation and BD Businesses. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product in research conducted by the buyer (whether the buyer is an academic or for-profit entity). The buyer cannot sell or otherwise transfer (a) this product (b) its components or (c) materials made using this product or its components to a third party or otherwise use this product or its components or materials made using this product or its components for Commercial Purposes. Commercial Purposes means any activity by a party for consideration and may include, but is not limited to: (1) use of the product or its components in manufacturing; (2) use of the product or its components to provide a service, information, or data; (3) use of the product or its components for therapeutic, diagnostic or prophylactic purposes; or (4) resale of the product or its components, whether or not such product or its components are resold for use in research. For information on purchasing a license to this product for any other use, contact Life Technologies Corporation, Cell Analysis Business Unit Business Development, 29851 Willow Creek Road, Eugene, OR 97402, USA, Tel: (541) 465-8300. Fax: (541) 335-0504.
      8. 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.
      9. When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
      10. Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
      11. Tandem fluorochromes contain both an energy donor and an energy acceptor. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from the donor may be observed. Additionally, multi-laser cytometers may directly excite both the donor and acceptor fluorochromes. Therefore, we recommend for every tandem conjugate, a matched individual single-stain control be acquired for generating a compensation or spectral unmixing matrix.
      12. CF™ is a trademark of Biotium, Inc.
      13. For U.S. patents that may apply, see bd.com/patents.
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      571245 Rev. 1
      Antibody Details
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      4a

      Stat (Signal transducer and activators of transcription) proteins mediate the biological activity of cytokines, including interleukins, interferons, erythropoietin, and growth factors.  Ligand-receptor interaction activates constitutively-associated JAK family kinases as well as subsequent recruitment and activation of Stat proteins by tyrosine phosphorylation.  Active Stat proteins then move to the nucleus to promote transcription of cytokine-inducible genes.  Seven Stat proteins have been cloned, each of which is differentially expressed and/or activated in a cytokine-specific and cell type-specific manner.  Stat1 and Stat2 are components of the ISGF3 (Interferon-Stimulated Gene Factor 3) complex, the primary transcription activator induced by interferon binding to a specific cell-surface receptor.  Stat1 has two alternatively spliced isoforms, 91-kDa Stat1α and 84-kDa Stat1β; Stat1α has 38 additional C-terminal amino acids.  In response to the binding of IFNα, IFNγ, EGF, PDGF, or CSF-1 to their respective receptors, the Stat1 subunits become tyrosine-phosphorylated at Y701, and the complex translocates to the nucleus. This forms an active complex that includes the DNA-binding p48 subunit, and is responsible for modulating interferon-stimulated genes (ISGs) transciption.

      The 4a monoclonal antibody recognizes the phosphorylated Y701 in Stat1α and Stat1β.

      571245 Rev. 1
      Format Details
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      RB613
      The BD Horizon RealBlue™ 613 (RB613) Dye is part of the BD® family of blue dyes. It is a tandem fluorochrome with an excitation maximum (Ex Max) at 492-nm and an emission maximum (Em Max) at 613-nm as measured using an antibody-dye conjugate. Driven by BD® innovation, RB613 can be used on both spectral and conventional cytometers and is designed to be excited by the Blue laser (488-nm) with reduced excitation by the 561-nm Yellow-Green laser. For conventional instruments equipped with a Blue laser (488-nm), RB613 can be used as an alternative to PE-CF594 and we recommend using an optical filter centered near 610-nm (eg, a 610/20-nm bandpass filter). For spectral instruments equipped with a Blue laser (488-nm), it can be used in conjunction with PE-CF594. RB613 is on average brighter than PE-CF594 off the blue laser.
      altImg
      RB613
      Blue 488 nm
      492 nm
      613 nm
      571245 Rev.1
      Citations & References
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      View product citations for antibody "571245" on CiteAb

      Development References (6)

      1. Bromberg J, Darnell JE. The role of STATs in transcriptional control and their impact on cellular function. Oncogene. 2000; 19(21):2468-2473. (Biology). View Reference
      2. Darnell JE Jr. STATs and gene regulation. Science. 1997; 277(5332):1630-1635. (Biology). View Reference
      3. Fu XY, Zhang JJ. Transcription factor p91 interacts with the epidermal growth factor receptor and mediates activation of the c-fos gene promoter. Cell. 1993; 74(6):1135-1145. (Biology). View Reference
      4. Perez OD, Mitchell D, Campos R, Gao GJ, Li L, Nolan GP. Multiparameter analysis of intracellular phosphoepitopes in immunophenotyped cell populations by flow cytometry. Curr Protoc Cytom. 2005; 6.20.1-6.20.22. (Clone-specific: Flow cytometry). View Reference
      5. Suni MA, Maino VC. Flow cytometric analysis of cell signaling proteins. Methods Mol Biol. 2011; 717:155-169. (Clone-specific: Flow cytometry). View Reference
      6. Tanaka S, Saito Y, Kunisawa J, et al. Development of mature and functional human myeloid subsets in hematopoietic stem cell-engrafted NOD/SCID/IL2rgammaKO mice. J Immunol. 2012; 188(12):6145-6155. (Clone-specific: Flow cytometry). View Reference
      View All (6) View Less
      571245 Rev. 1

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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.