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RB670 Mouse Anti-Human Granzyme B
RB670 Mouse Anti-Human Granzyme B
Two-color flow cytometric analysis of Granzyme B expression in Human peripheral blood lymphocytes.  Human whole blood was fixed with BD Lyse/Fix Buffer (Cat. No. 558049). The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with BD Horizon™ BUV395 Mouse Anti-Human CD8 antibody (Cat. No. 563795) and with either BD Horizon™ RB670 Mouse IgG1, κ Isotype Control (Cat. No. 571784; Left Plot) or BD Horizon™ RB670 Mouse Anti-Human Granzyme B antibody (Cat. No. 571919/571920; Right Plot). The bivariate pseudocolor density plot showing the correlated expression of Granzyme B (or Ig Isotype control staining) versus CD8 was derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer and FlowJo™ Software..
Two-color flow cytometric analysis of Granzyme B expression in Human peripheral blood lymphocytes.  Human whole blood was fixed with BD Lyse/Fix Buffer (Cat. No. 558049). The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with BD Horizon™ BUV395 Mouse Anti-Human CD8 antibody (Cat. No. 563795) and with either BD Horizon™ RB670 Mouse IgG1, κ Isotype Control (Cat. No. 571784; Left Plot) or BD Horizon™ RB670 Mouse Anti-Human Granzyme B antibody (Cat. No. 571919/571920; Right Plot). The bivariate pseudocolor density plot showing the correlated expression of Granzyme B (or Ig Isotype control staining) versus CD8 was derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer and FlowJo™ Software..
Product Details
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BD Horizon™
GZMB; Granzyme-2; CCPI; CGL1; CSPB; CTLA1; CTSGL1; GRB; HLP; SECT
Human (QC Testing)
Mouse BALB/c IgG1, κ
Human Granzyme B
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl/test
3002
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.

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.

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. 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).
  4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. An isotype control should be used at the same concentration as the antibody of interest.
  6. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  7. Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
  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. For U.S. patents that may apply, see bd.com/patents.
  10. 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.
  11. 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.
  12. 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.
571920 Rev. 1
Antibody Details
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GB11

The GB11 antibody specifically reacts with human granzyme B, a serine protease of  approximately 32 kDa.  Granzyme B is stored in the granules of cytotoxic T lymphocytes and NK cells along with the pore-forming protein perforin. In the classic model of target cell lysis, perforins create holes in the target cell membrane allowing entrance of granzymes.  Granzyme B has been shown to act on specific substrates including caspase-3, -7, -9, and -10 which in turn give rise to enzymes that mediate apoptosis. Granzyme B may also be involved in the hydrolysis of extracellular matrix components.  Detectable levels of granzyme B have been detected in sera from healthy volunteers. The immunogen used to generate the GB11 hybridoma was human granzyme B isolated from an NK cell line.

571920 Rev. 1
Format Details
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RB670
The BD Horizon RealBlue™ 670 (RB670) 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 670 nm as measured using an antibody-dye conjugate. Driven by BD® innovation, RB670 can be used on both spectral and conventional cytometers and is designed to be primarily excited by the Blue laser (488-nm). For conventional instruments equipped with only a Blue laser (488-nm), RB670 can be used as an alternative to PE-Cy5 and we recommend using an optical filter centered near 670-nm (eg, a 670/30-nm bandpass filter). For conventional and spectral instruments equipped with both a Blue (488-nm) and Yellow-Green (561-nm) laser and appropriate detectors, it can be used in conjunction with PE-Cy5.
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RB670
Blue 488 nm
492 nm
670 nm
571920 Rev.1
Citations & References
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View product citations for antibody "571920" on CiteAb

Development References (8)

  1. Hamann D, Baars PA, Rep MH. Phenotypic and functional separation of memory and effector human CD8+ T cells. J Exp Med. 1997; 186(9):1407-1418. (Clone-specific: Intracellular Staining/Flow Cytometry). View Reference
  2. Poe M, Blake JT, Boulton DA. Human cytotoxic lymphocyte granzyme B. Its purification from granules and the characterization of substrate and inhibitor specificity. J Biol Chem. 1991; 266(1):98-103. (Biology). View Reference
  3. Ronday HK, van der Laan WH, Tak PP et al. Human granzyme B mediates cartilage proteoglycan degradation and is expressed at the invasive front of the synovium in rheumatoid arthritis. Rheumatology (Oxford). 2001; 40:55-61. (Biology). View Reference
  4. Smyth MJ, Kelly JM, Sutton VR et al. Unlocking the secrets of cytotoxic granule proteins. J Leukoc Biol. 2001; 70:18-29. (Biology). View Reference
  5. Spaeny-Dekking EH, Hanna WL, Wolbink AM et al. Extracellular granzymes A and B in humans: detection of native species during CTL responses in vitro and in vivo. J Immunol. 1998; 160:3610. (Immunogen: ELISA, Radioimmunoassay). View Reference
  6. Trapani JA, Klein JL, White PC, and Dupont B. Molecular cloning of an inducible serine esterase gene from human cytotoxic lymphocytes. Proc Natl Acad Sci U S A. 1988; 5:6924-6928. (Biology). View Reference
  7. Trapani JA, Smyth MJ, Apostolidis VA, Dawson M, and Browne KA. Granule serine proteases are normal nuclear constituents of natural killer cells. J Biol Chem. 1994; 269:18359-18365. (Biology). View Reference
  8. Wever PC, Van Der Vliet HJ, Spaeny LH . The CD8+ granzyme B+ T-cell subset in peripheral blood from healthy individuals contains activated and apoptosis-prone cells. Immunology. 1998; 93(3):383-389. (Immunogen: ELISA, Flow cytometry, Immunoprecipitation, Intracellular Staining/Flow Cytometry, Radioimmunoassay). View Reference
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571920 Rev. 1

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