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Alexa Fluor® 488 Mouse anti-Bcl-6
Alexa Fluor® 488 Mouse anti-Bcl-6
  Bcl-6 expression in Human B lymphocytes (Left Panel). Tonsil cells were stained with BD Horizon™ V450 Anti-Human CD19 (Cat. No. 560353), PE-Cy™7 Anti-Human CD4 (Cat. No. 560649) and APC Anti-Human CD38 (Cat. No. 555462). Cells were washed, fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885), followed by no staining (Left Figure) or intracellular staining with Alexa Fluor® 488 Anti-Bcl-6 (Right Figure; Cat. No. 561524). Contour plots show expression of Bcl-6 (or background fluorescence) versus CD38 by B cells identified as CD4-CD19+ events with the light-scatter characteristics of intact lymphocytes. Bcl-6 staining is observed in CD38+ germinal center B cells but not in CD38- naïve B cells nor  CD38++ plasma cells. Cytometry was performed with a BD FACSCanto™ II Flow Cytometer. Data courtesy of Mark Kroenke & Shane Crotty, La Jolla Institute for Allergy & Immunology.   Bcl-6 expression in Mouse B lymphocytes (Right Panel). BALB/c mesenteric lymph node cells were stained with APC Anti-Mouse B220 (Cat. No. 553092), PerCP-Cy™5.5 Anti-Mouse CD4 (Cat. No. 550954), PE Anti-Mouse Fas/CD95 (Cat. No. 554258), and BD Horizon™ V450 Anti-Mouse IgD (Cat. No. 560869). Cells were washed, resuspended in RPMI with 10% FBS, and fixed with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049). Cells were permeabilized with BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885), followed by intracellular staining with Alexa Fluor® 488 Anti-Bcl-6 (Cat. No. 561524). A contour plot shows the expression of IgD versus Fas/CD95 by B cells identified as CD4-B220+ events with the light-scatter characteristics of intact lymphocytes (Left Figure). Germinal center B cells were identified as IgDloFas+ B lymphocytes. Flow histograms (Right Figure) show intracellular Bcl-6 staining of mouse germinal center B cells (open histogram) and non-GC B cells (shaded histogram). Cytometry was performed with a BD LSRFortessa™ Flow Cytometer.
  Bcl-6 expression in Human B lymphocytes (Left Panel). Tonsil cells were stained with BD Horizon™ V450 Anti-Human CD19 (Cat. No. 560353), PE-Cy™7 Anti-Human CD4 (Cat. No. 560649) and APC Anti-Human CD38 (Cat. No. 555462). Cells were washed, fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885), followed by no staining (Left Figure) or intracellular staining with Alexa Fluor® 488 Anti-Bcl-6 (Right Figure; Cat. No. 561524). Contour plots show expression of Bcl-6 (or background fluorescence) versus CD38 by B cells identified as CD4-CD19+ events with the light-scatter characteristics of intact lymphocytes. Bcl-6 staining is observed in CD38+ germinal center B cells but not in CD38- naïve B cells nor  CD38++ plasma cells. Cytometry was performed with a BD FACSCanto™ II Flow Cytometer. Data courtesy of Mark Kroenke & Shane Crotty, La Jolla Institute for Allergy & Immunology.   Bcl-6 expression in Mouse B lymphocytes (Right Panel). BALB/c mesenteric lymph node cells were stained with APC Anti-Mouse B220 (Cat. No. 553092), PerCP-Cy™5.5 Anti-Mouse CD4 (Cat. No. 550954), PE Anti-Mouse Fas/CD95 (Cat. No. 554258), and BD Horizon™ V450 Anti-Mouse IgD (Cat. No. 560869). Cells were washed, resuspended in RPMI with 10% FBS, and fixed with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049). Cells were permeabilized with BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885), followed by intracellular staining with Alexa Fluor® 488 Anti-Bcl-6 (Cat. No. 561524). A contour plot shows the expression of IgD versus Fas/CD95 by B cells identified as CD4-B220+ events with the light-scatter characteristics of intact lymphocytes (Left Figure). Germinal center B cells were identified as IgDloFas+ B lymphocytes. Flow histograms (Right Figure) show intracellular Bcl-6 staining of mouse germinal center B cells (open histogram) and non-GC B cells (shaded histogram). Cytometry was performed with a BD LSRFortessa™ Flow Cytometer.
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BD Pharmingen™
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_10716202
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 Alexa Fluor® 488 under optimum conditions, and unreacted Alexa Fluor® 488 was removed.

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This fluorescent antibody conjugate is suitable for intracellular staining of cell lines and primary cells fixed using BD Cytofix™ Fixation Buffer (Cat. No. 554655) or BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) and permeabilized using BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885) or BD Phosflow™ Perm Buffers II (Cat. No. 558052), III (Cat. No. 558050) or IV (Cat. No. 560746).

We validate the quality of each batch of the K112-91 antibody conjugate by flow cytometry on human cell lines.  Investigators may use the same cell lines as controls for their staining procedure, namely Ramos (Positive; ATCC CRL-1596) and Jurkat (Negative; ATCC TIB-152) human cell lines actively growing in log phase (do not overgrow).  Cells are fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655; 10 minutes at 37°C), permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050; 30 minutes on ice), and washed using BD Pharmingen™ Stain Buffer (Cat. No. 554656), followed by intracellular staining with Mouse anti-Bcl-6 for 45 minutes at room temperature.

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. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  5. Alexa Fluor® 488 fluorochrome emission is collected at the same instrument settings as for fluorescein isothiocyanate (FITC).
  6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  7. The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
  8. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
  9. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  10. Cy is a trademark of GE Healthcare.
  11. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
561524 Rev. 5
抗体の詳細
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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 K112-91 antibody is compatible with multiple cellular fixation and permeabilization treatments as listed in the Recommended Assay Procedure section (please see below).

             

561524 Rev. 5
フォーマットの詳細
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Alexa Fluor™ 488
Alexa Fluor™ 488 Dye is part of the BD blue family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 494-nm and an emission maximum (Em Max) at 517-nm. Alexa Fluor™ 488 is designed to be excited by the Blue laser (488 nm) and detected using an optical filter centered near 520-nm (e.g., a 530/30-nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
Alexa Fluor™ 488
Blue 488 nm
494 nm
517 nm
561524 Rev.5
引用&参考文献
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Development References (9)

  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. 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). View Reference
  3. 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
  4. Crotty S. Follicular Helper CD4 T Cells (Tfh). Annu Rev Immunol. 2011; 29(1):621-663. (Biology). View Reference
  5. 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
  6. 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
  7. 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
  8. 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
  9. Ye BH, Lista F, Lo Coco F, et al. Alterations of a zinc finger-encoding gene, BCL-6, in diffuse large-cell lymphoma. Science. 1993; 262(5134):747-750. (Biology). View Reference
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561524 Rev. 5

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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.