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Alexa Fluor® 647 Mouse IgG1 κ Isotype Control
Alexa Fluor® 647 Mouse IgG1 κ Isotype Control

Expression of TNF by stimulated human peripheral blood lymphocytes. Human peripheral blood mononuclear cells were stimulated for 4 hours with PMA (5 ng/ml, Sigma, Cat. No. P-8139) in the presence of Brefeldin A (BD GolgiPlug™ Protein Transport Inhibitor, Cat. No. 555029). Cells were harvested, fixed, permeabilized and stained with PE Mouse Anti-Human CD8 (Cat. No. 555367) and either Alexa Fluor®647 Mouse Anti Human TNF (Cat. No. 557733; left panel) or Alexa Fluor® 647 MOPC-21 immunoglobulin (Cat. No. 557732, right panel). Dot plots were derived from gated events with the forward and side light scatter characteristics of lymphocytes. The quadrant markers for the bivariate dot plots were set based on the autofluorescence and isotype controls.

Expression of TNF by stimulated human peripheral blood lymphocytes. Human peripheral blood mononuclear cells were stimulated for 4 hours with PMA (5 ng/ml, Sigma, Cat. No. P-8139) in the presence of Brefeldin A (BD GolgiPlug™ Protein Transport Inhibitor, Cat. No. 555029). Cells were harvested, fixed, permeabilized and stained with PE Mouse Anti-Human CD8 (Cat. No. 555367) and either Alexa Fluor®647 Mouse Anti Human TNF (Cat. No. 557733; left panel) or Alexa Fluor® 647 MOPC-21 immunoglobulin (Cat. No. 557732, right panel). Dot plots were derived from gated events with the forward and side light scatter characteristics of lymphocytes. The quadrant markers for the bivariate dot plots were set based on the autofluorescence and isotype controls.

Product Details
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BD Pharmingen™
Mouse IgG1, κ
Intracellular staining (flow cytometry), Isotype control (Routinely Tested)
5 µl
AB_396840
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

Intracellular Immunofluorescent Staining Analysis: The MOPC-21 immunoglobulins are suitable mouse IgG1, κ isotype controls for assessing the level of background staining on paraformaldehyde fixed/saponin-permeabilized mouse or human cells for flow cytometric analysis. Use the MOPC-21 antibody at 5 µl test as recommended to serve as an isotype control.

BD Pharmingen™ Alexa Fluor® 647 Mouse IgG1 κ Isotype control, Cat. No. 557714 is recommended for non-intracellular staining.

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. 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.
  6. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
  7. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  8. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
  9. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
557732 Rev. 4
Antibody Details
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MOPC-21

The MOPC-21 immunoglobulin is a mouse myeloma protein. The MOPC-21 immunoglobulin was selected as an isotype control following screening for low background on a variety of mouse and human tissues.

557732 Rev. 4
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
557732 Rev.4
Citations & References
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Development References (1)

  1. Prussin C, Metcalfe DD. Detection of intracytoplasmic cytokine using flow cytometry and directly conjugated anti-cytokine antibodies. J Immunol Methods. 1995; 188(1):117-128. (Clone-specific: Blocking, Flow cytometry). View Reference
557732 Rev. 4

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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.