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R718 Mouse Anti-Bcl-6
R718 Mouse Anti-Bcl-6

Flow cytometric analysis of Bcl-6 expression in Ramos cells. Cells from the human Ramos (Burkitt's lymphoma, ATCC CRL-1596) cell line were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050) followed by intracellular staining with either BD Horizon™ R718 Mouse IgG1, κ Isotype Control (Cat. No. 566928; dashed line histogram) or BD Horizon™ R718 Mouse Anti-Human Bcl-6 antibody (Cat. No. 566979/566993; solid line histogram). A fluorescence histogram showing Bcl-6 expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of intact cells. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.

Flow cytometric analysis of Bcl-6 expression in Ramos cells. Cells from the human Ramos (Burkitt's lymphoma, ATCC CRL-1596) cell line were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050) followed by intracellular staining with either BD Horizon™ R718 Mouse IgG1, κ Isotype Control (Cat. No. 566928; dashed line histogram) or BD Horizon™ R718 Mouse Anti-Human Bcl-6 antibody (Cat. No. 566979/566993; solid line histogram). A fluorescence histogram showing Bcl-6 expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of intact cells. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.

Product Details
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BD Horizon™
BCL6; B-cell lymphoma 6 protein; LAZ3; Laz-3, ZBTB27, ZNF51
Human (QC Testing), Mouse (Tested in Development)
Mouse IgG1, κ
Human Bcl-6 Recombinant Protein
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_2869985
Aqueous buffered solution containing ≤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 unreacted dye was removed.

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. 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. 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.
  4. Alexa Fluor® is a registered trademark of Life Technologies Corporation.
  5. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  6. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  7. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  8. This product is provided under an Agreement between BIOTIUM and BD Biosciences. This product, and only in the amount purchased by buyer, may be used solely for buyer’s own internal research, in a manner consistent with the accompanying product literature. No other right to use, sell or otherwise transfer (a) this product, or (b) its components is hereby granted expressly, by implication or by estoppel. This product is for research use only. Diagnostic uses require a separate license from Biotium, Inc. For information on purchasing a license to this product including for purposes other than research, contact Biotium, Inc., 3159 Corporate Place, Hayward, CA 94545, Tel: (510) 265-1027. Fax: (510) 265-1352. Email: btinfo@biotium.com.
  9. Alexa Fluor™ is a trademark of Life Technologies Corporation.
566979 Rev. 5
Antibody Details
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K112-91

The K112-91 monoclonal antibody specifically binds to Bcl-6. Bcl-6 was first identified as a proto-oncogene frequently deregulated by chromosomal translocations in non-Hodgkin B-cell lymphomas. It is a nuclear transcriptional repressor of the BTB/POZ zinc-finger family of transcription factors. In addition to its roles in cancer, Bcl-6 plays important roles in the differentiation of normal cells including B cells, thymocytes, CD4+ or CD8+ T cells. Bcl-6 is highly expressed in germinal center B cells, where it promotes the germinal center reaction by inducing proliferation and inhibiting the DNA-damage response. Bcl-6 has been identified as a key factor in promoting the differentiation of CD4+ follicular T helper (Tfh) cells that are involved in promoting germinal center formation and providing help to B cells. The interplay of Bcl-6 and another transcriptional repressor, Blimp-1, is thought to be critical in defining the results of both B-cell and T-cell differentiation.

The antibody was conjugated to BD Horizon™ Red 718, which has been developed exclusively by for BD Biosciences as a better alternative to Alexa Fluor™ 700. BD Horizon™ Red 718 can be excited by the red laser (628 – 640 nm) and, with an Em Max around 718 nm, it can be detected using a 730/45 nm filter. Due to similar excitation and emission properties, we do not recommend using R718 in combination with APC-R700 or Alexa Fluor™ 700.

566979 Rev. 5
Format Details
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R718
The BD Horizon™ Red 718 (R718) Dye is part of the BD red family of dyes. It is a small organic fluorochrome with an excitation maximum (Ex Max) at 695-nm and an emission maximum (Em Max) at 718-nm. Driven by BD innovation, R718 is designed to be excited by the red laser (627–640-nm) and detected using an optical filter centered near 720-nm (e.g., a 720/40-nm bandpass filter). R718 is a brighter alternative to Alexa Fluor™ 700. R718 is also a bright small molecule alternative to APC-R700 with lower spread into the APC detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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R718
Red 627-640 nm
695 nm
718 nm
566979 Rev.5
Citations & References
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Development References (10)

  1. Baumjohann D, Okada T, Ansel KM. Cutting Edge: Distinct Waves of BCL6 Expression during T Follicular Helper Cell Development. J Immunol. 2011; 187(5):2089-2092. (Clone-specific: Flow cytometry). View Reference
  2. Chung Y, Tanaka S, Chu F, et al. Follicular regulatory T cells expressing Foxp3 and Bcl-6 suppress germinal center reactions. Nat Med. 2011; 17(8):983-988. (Clone-specific: Flow cytometry). View Reference
  3. Crotty S, Choi YS, Kageyama R, et al. ICOS receptor instructs T follicular helper cell versus effector cell differentiation via induction of the transcriptional repressor Bcl6. Immunity. 2011; 34:1-15. (Clone-specific: Flow cytometry). View Reference
  4. Crotty S, Johnston RJ, Schoenberger SP. Effectors and memories: Bcl-6 and Blimp-1 in T and B lymphocyte differentiation. Nat Immunol. 2010; 11(2):114-120. (Biology). View Reference
  5. Crotty S. Follicular Helper CD4 T Cells (Tfh). Annu Rev Immunol. 2011; 29(1):621-663. (Biology). View Reference
  6. Eto, D., C. Lao, et al. IL-21 and IL-6 are critical for different aspects of B cell immunity and redundantly induce optimal follicular helper CD4 T cell (Tfh) differentiation. PLoS ONE. 2011; 6(3):e17739. (Clone-specific: Flow cytometry). View Reference
  7. Fazilleau N, McHeyzer-Williams LJ, Rosen H, McHeyzer-Williams MG. The function of follicular helper T cells is regulated by the strength of T cell antigen receptor binding. Nat Rev Immunol. 2009; 10(4):375-384. (Biology). View Reference
  8. Johnston RJ, Poholek AC, DiToro D, et al. Bcl6 and Blimp-1 are reciprocal and antagonistic regulators of T follicular helper cell differentiation.. Science. 2009; 325(5943):1006-10. (Biology). View Reference
  9. Klein U, Dalla-Favera R. Germinal centres: role in B-cell physiology and malignancy. Nat Rev Immunol. 2008; 8(1):22-33. (Biology). View Reference
  10. Nurieva RI, Chung Y, Martinez GJ, et al. Bcl6 mediates the development of T follicular helper cells. Science. 2009; 325(5943):1001-1005. (Biology). View Reference
View All (10) View Less
566979 Rev. 5

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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.