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Alexa Fluor® 647 Mouse Anti-Human CD20
Alexa Fluor® 647 Mouse Anti-Human CD20
Analysis of CD20 (cytoplasmic) in human peripheral blood lymphocytes.  Human peripheral blood mononuclear cells were fixed with pre-warmed BD Cytofix™ buffer (Cat. No. 554655) for 10 minutes at 37ºC and permeabilized with BD Phosflow™ Perm Buffer II (Cat. No. 558052) on ice for 30 minutes.  The cells were then stained with either Alexa Fluor® 647 Mouse IgG2a, κ, isotype control(Cat. No. 558053, thin line) or Alexa Fluor® 647 Mouse anti-human CD20 (cytoplasmic) (bold line).  For data analysis, lymphocytes were selected by their scatter profile.  CD20 (cytoplasmic) was not detected in the monocytes (not shown).  Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.
Analysis of CD20 (cytoplasmic) in human peripheral blood lymphocytes.  Human peripheral blood mononuclear cells were fixed with pre-warmed BD Cytofix™ buffer (Cat. No. 554655) for 10 minutes at 37ºC and permeabilized with BD Phosflow™ Perm Buffer II (Cat. No. 558052) on ice for 30 minutes.  The cells were then stained with either Alexa Fluor® 647 Mouse IgG2a, κ, isotype control(Cat. No. 558053, thin line) or Alexa Fluor® 647 Mouse anti-human CD20 (cytoplasmic) (bold line).  For data analysis, lymphocytes were selected by their scatter profile.  CD20 (cytoplasmic) was not detected in the monocytes (not shown).  Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.
Product Details
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BD Phosflow™
MS4A1; membrane-spanning 4-domains subfamily A member 1; B1; Bp35; LEU-16
Human (QC Testing)
Mouse BALB/c IgG2a, κ
Human B lymphoma cell line
Intracellular staining (flow cytometry) (Routinely Tested)
20 µl
V cB010
AB_647118
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® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed.

Recommended Assay Procedures

This antibody conjugate is suitable for intracellular staining of human whole blood (using BD Phosflow™ Lyse/Fix Buffer) and peripheral blood mononuclear cells (using BD Cytofix™ Fixation Buffer or BD Phosflow™ Fix Buffer I).  Any of the three BD Phosflow™ permeabilization buffers may be used.

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. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  3. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  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. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  6. 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.
  7. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
  8. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
  9. An isotype control should be used at the same concentration as the antibody of interest.
558054 Rev. 8
Antibody Details
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H1

The H1 (FB1) antibody specificially binds to a cytoplasmic domain of CD20. CD20 is a 33-37-kDa four transmembrane phosphoprotein that is expressed by B lymphocytes from the pre-B stage and most malignant B cells and is lost during plasma cell differentiation.  Low level CD20 expression is observed on a subset of normal circulating T lymphocytes, and CD20-positive T-cell lymphomas have been reported.  The CD20 molecule is associated with membrane lipid raft domains, acts as a channel for calcium ions, and is involved in the regulation of B cell activation and survival.  The cytoplasmic domain regions are serine and threonine rich and contain multiple phosphorylation consensus sequences.

558054 Rev. 8
Format Details
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Alexa Fluor™ 647
Alexa Fluor™ 647 Dye is part of the BD red family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 653-nm and an emission maximum (Em Max) at 669-nm. Alexa Fluor 647 is designed to be excited by the Red laser (627-640 nm) and detected using an optical filter centered near 520-nm (e.g., a 660/20 nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
Alexa Fluor™ 647
Red 627-640 nm
653 nm
669 nm
558054 Rev.8
Citations & References
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Development References (4)

  1. Cragg MS, Walshe CA, Ivanov AO, Glennie MJ. The biology of CD20 and its potential as a target for mAb therapy. Curr Dir Autoimmun. 2005; 8:140-174. (Biology). View Reference
  2. Kitamura A, Yamashita Y, Mori N. CD20-positive cytotoxic T cell lymphoma: report of two cases and review of the literature. J Clin Exp Hematop. 2005; 45(1):45-50. (Biology).
  3. Nozawa Y, Abe M, Ohno H, Fukuhara S, Wakasa H. Production of two monoclonal antibodies (FB1 and FB21) useful for the identification of human B lymphocytes in formalin-fixed, paraffin-embedded tissues. J Pathol. 1994; 173:347-354. (Immunogen). View Reference
  4. Nozawa Y, Abe M, Wakasa H. Three mAb, FUN-1, FB1, and FB21, that recognize B-cell antigens in frozen or paraffin-embedded tissue sections. In: Schlossman SF. Stuart F. Schlossman .. et al., ed. Leucocyte typing V : white cell differentiation antigens : proceedings of the fifth international workshop and conference held in Boston, USA, 3-7 November, 1993. Oxford: Oxford University Press; 1995:705-706.
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558054 Rev. 8

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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.