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BB515 Mouse Anti-Human CD54
BB515 Mouse Anti-Human CD54

Two-parameter flow cytometric analysis of CD54 expression on human peripheral blood leucocytes. Human whole blood was stained with either BD Horizon™ BB515 Mouse IgG1, κ Isotype Control (Cat. No. 564416; Left Panel) or BD Horizon BB515 Mouse Anti-Human CD54 antibody (Cat. No. 564685; Right Panel). Erythrocytes were lysed with BD FACS Lysing Solution (Cat. No. 349202).  Two-parameter flow cytometric contour plots showing the correlated expression of CD54 (or Ig Isotype control staining) versus side light-scatter signals were derived from gated events with the forward and side light-scatter characteristics of intact leucocyte populations. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

Two-parameter flow cytometric analysis of CD54 expression on human peripheral blood leucocytes. Human whole blood was stained with either BD Horizon™ BB515 Mouse IgG1, κ Isotype Control (Cat. No. 564416; Left Panel) or BD Horizon BB515 Mouse Anti-Human CD54 antibody (Cat. No. 564685; Right Panel). Erythrocytes were lysed with BD FACS Lysing Solution (Cat. No. 349202).  Two-parameter flow cytometric contour plots showing the correlated expression of CD54 (or Ig Isotype control staining) versus side light-scatter signals were derived from gated events with the forward and side light-scatter characteristics of intact leucocyte populations. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.

Product Details
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BD Horizon™
ICAM-1; intercellular adhesion molecule 1; BB2; P3.58
Human (QC Testing)
Mouse BALB/c IgG1, κ
IFN-γ-treated BM314 colonic carcinoma cells
Flow cytometry (Routinely Tested)
5 µl
VI A095
3383
AB_2738892
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. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  6. 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.
  7. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
564685 Rev. 2
Antibody Details
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HA58

The HA58 monoclonal antibody specifically binds to CD54 which is also known as ICAM-1 (intracellular adhesion molecule-1).  CD54 is an 85-110 kDa type I transmembrane glycoprotein that belongs to the immunoglobulin supergene family. A soluble form of CD54 can also be found in biological fluids. CD54 is expressed on endothelial cells and both resting (weak) and activated (moderate) lymphocytes and monocytes. CD54 is a ligand for LFA-1 (CD11a/CD18) and Mac-1 (CD11b/CD18). The CD54 adhesion molecule plays roles in inflammatory and immune responses and neoplasia. This antibody (NA/LE format) blocks the mixed-lymphocyte reaction (MLR) and the purified format is suitable for staining acetone-fixed, frozen tissue sections.

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.

564685 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
564685 Rev.2
Citations & References
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Development References (4)

  1. Kishimoto T. Tadamitsu Kishimoto .. et al., ed. Leucocyte typing VI : white cell differentiation antigens : proceedings of the sixth international workshop and conference held in Kobe, Japan, 10-14 November 1996. New York: Garland Pub.; 1997.
  2. Knapp W. W. Knapp .. et al., ed. Leucocyte typing IV : white cell differentiation antigens. Oxford New York: Oxford University Press; 1989:1-1182.
  3. 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.
  4. Tsujisaki M, Imai K, Hirata H, et al. Detection of circulating intercellular adhesion molecule-1 antigen in malignant diseases. Clin Exp Immunol. 1991; 85(1):3-8. (Immunogen: Blocking, ELISA, Flow cytometry, Immunoprecipitation, Radioimmunoassay, Western blot). View Reference
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564685 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.