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BB515 Mouse Anti-Human CD15
BB515 Mouse Anti-Human CD15

Flow cytometric analysis of CD15 expression on human peripheral blood granulocytes - Staining comparisons between BD Horizon™ BB515- and FITC-conjugated antibodies. Whole blood was stained with BD Horizon BB515 Mouse IgM, κ Isotype Control (Cat. No. 564680; dashed line histogram) or BD Horizon BB515 Mouse Anti-Human CD15 antibody (Cat. No. 565236; bold solid line histograms). Alternatively, cells were stained with FITC Anti-Human CD15 antibody (Cat. No. 555401/ 560997; thin solid line histogram). The erythrocytes were lysed with BD FACS™ Lysing Solution (Cat. No. 349202).

       Overlaid histograms are shown to facilitate staining comparisons between: BB515 Anti-CD15 antibody versus its Ig Isotype Control (Left Panel), and BB515 Anti-CD15 antibody versus FITC Anti-CD15 antibody (Right Panel). The fluorescence histograms showing CD15 expression (or Ig Isotype control staining) were derived from gated events with the forward and side light-scatter characteristics of intact granulocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

Flow cytometric analysis of CD15 expression on human peripheral blood granulocytes - Staining comparisons between BD Horizon™ BB515- and FITC-conjugated antibodies. Whole blood was stained with BD Horizon BB515 Mouse IgM, κ Isotype Control (Cat. No. 564680; dashed line histogram) or BD Horizon BB515 Mouse Anti-Human CD15 antibody (Cat. No. 565236; bold solid line histograms). Alternatively, cells were stained with FITC Anti-Human CD15 antibody (Cat. No. 555401/ 560997; thin solid line histogram). The erythrocytes were lysed with BD FACS™ Lysing Solution (Cat. No. 349202).

       Overlaid histograms are shown to facilitate staining comparisons between: BB515 Anti-CD15 antibody versus its Ig Isotype Control (Left Panel), and BB515 Anti-CD15 antibody versus FITC Anti-CD15 antibody (Right Panel). The fluorescence histograms showing CD15 expression (or Ig Isotype control staining) were derived from gated events with the forward and side light-scatter characteristics of intact granulocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

Product Details
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BD Horizon™
Lewis X; Le-X; X-Hapten; SSEA-1; 3-FAL
Human (QC Testing)
Mouse IgM, κ
Human Mononuclear Cells
Flow cytometry (Routinely Tested)
5 µl
IV M141
AB_2739127
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 with BD Horizon™ BB515 under optimum conditions and unconjugated antibody was removed.

Recommended Assay Procedures

BD™ CompBeads can be used as surrogates to assess fluorescence spillover (Compensation).  When fluorochrome conjugated antibodies are bound to 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 CompBead to ensure that BD Comp beads are appropriate for your specific cellular application.

For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes are used in the same experiment.  Fluorescent dye interactions may cause staining artifacts which may affect data interpretation.  The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions.  More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).

For optimal results, it is recommended to perform 2 washes after staining with antibodies. Cells may be prepared, stained with antibodies and washed twice with wash buffer per established protocols for immunofluorescence staining, prior to acquisition on a flow cytometer. Performing fewer than the recommended wash steps may lead to increased spread of the negative population.

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. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. BD Horizon Brilliant Stain Buffer is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,575,303; 8,354,239.
  6. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  7. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
565236 Rev. 2
Antibody Details
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HI98

The HI98 monoclonal antibody specifically reacts with 3-fucosyl-N-acetyllactosamine (3-FAL), a 220 kDa carbohydrate structure, also called X-hapten, SSEA-1, CD15 or Lewis X. This structure is found on a variety of cell surface glycolipids and glycoproteins. 3-FAL is expressed on >95% of granulocytes, including neutrophils and eosinophils, and to a varying degree on monocytes, but not on lymphocytes or basophils. CD15 plays a role in mediating phagocytosis, bactericidal activity and chemotaxis. This antibody is also suitable for staining formalin-fixed, paraffin-embedded tissue sections without pretreatment. Since the Abs are recognizing a carbohydrate epitope (3-fucosyl-N-acetyllactosamine) they also should work across species and not only for human.

The antibody was conjugated to BD Horizon BB515 which is part of the BD Horizon Brilliant™ Blue family of dyes. With an Ex Max near 490 nm and an Em Max near 515 nm, BD Horizon BB515 can be excited by the blue laser (488 nm) laser and detected with a 530/30 nm filter. This dye has been exclusively developed by BD Biosciences and is up to seven times brighter than FITC with less spillover into the PE channel. Due to similar excitation and emission properties, BB515, FITC, and Alexa Fluor® 488 cannot be used simultaneously. It is not recommended to use BB515 in cocktails that include Streptavidin conjugates as it may cause high background.

565236 Rev. 2
Format Details
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BB515
The BD Horizon Brilliant™ Blue 515 (BB515) dye is part of the BD Horizon Brilliant™ Blue family of dyes. This dye is a polymer fluorochrome with an excitation maximum (Ex Max) at 490-nm and an emission maximum (Em Max) of 515-nm. Driven by BD innovation, BB515 is designed to be excited by the blue laser (488-nm) and detected using an optical filter centered near 520-nm (e.g., 530/30-nm). BB515 reagents are significantly brighter than equivalent FITC or Alexa Fluor™ 488 reagents with less spillover into the PE detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BB515
Blue 488 nm
490 nm
515 nm
565236 Rev.2
Citations & References
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Development References (7)

  1. Emanuel PD, Peiper SC, Chen Z, Sheng DC, Zuckerman KS. Specific inhibition of interleukin 3 bioactivity by a monoclonal antibody reactive with hematopoietic progenitor cells. Proc Natl Acad Sci U S A. 1990; 87(12):4449-4452. (Immunogen: Functional assay, Inhibition). View Reference
  2. Knapp W. Flow cytometry studies with blind panel antibodies. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:851-854.
  3. Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:1-1182.
  4. Koller U. Summary of immunohistology studies. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:862-867.
  5. Lund-Johansen F, Olweus J, Horejsi V, et al. Activation of human phagocytes through carbohydrate antigens (CD15, sialyl-CD15, CDw17, and CDw65).. J Immunol. 1992; 148(10):3221-9. (Biology). View Reference
  6. Shah VO, Safford MG, Terstappen LWMM, Loken MR. Quantitative comparison of myeloid antigens on peripheral blood lymphocytes, monocytes, neutrophils, eosinophils, and basophils. In: Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:855-858.
  7. Zola H. Leukocyte and stromal cell molecules : the CD markers. Hoboken, N.J.: Wiley-Liss; 2007.
View All (7) View Less
565236 Rev. 2

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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.