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BB515 Rat Anti-Mouse CD45
BB515 Rat Anti-Mouse CD45

Flow cytometric analysis of CD45 expression on mouse splenocytes - Staining comparisons between BD Horizon™ BB515- and FITC-conjugated antibodies. Mouse splenic leucocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). The cells were then stained with either BD Horizon™ BB515 Rat IgG2b, κ Isotype Control (Cat. No. 564421; dashed line histogram) or BD Horizon BB515 Rat Anti-Mouse CD45 antibody (Cat. No. 564590; bold solid line histogram). Alternatively, cells were stained with FITC Anti-Mouse CD45 antibody (Cat. No. 553079/553080/561088; thin solid line histogram).

       Overlaid histograms are shown to facilitate staining comparisons between: BB515 Anti-CD45 antibody versus its Ig Isotype Control (Left Panel), and BB515 Anti-CD45 antibody versus FITC Anti-CD45 antibody (Right Panel). The fluorescence histograms showing CD45 expression (or Ig Isotype control staining) were derived from gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.

Flow cytometric analysis of CD45 expression on mouse splenocytes - Staining comparisons between BD Horizon™ BB515- and FITC-conjugated antibodies. Mouse splenic leucocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). The cells were then stained with either BD Horizon™ BB515 Rat IgG2b, κ Isotype Control (Cat. No. 564421; dashed line histogram) or BD Horizon BB515 Rat Anti-Mouse CD45 antibody (Cat. No. 564590; bold solid line histogram). Alternatively, cells were stained with FITC Anti-Mouse CD45 antibody (Cat. No. 553079/553080/561088; thin solid line histogram).

       Overlaid histograms are shown to facilitate staining comparisons between: BB515 Anti-CD45 antibody versus its Ig Isotype Control (Left Panel), and BB515 Anti-CD45 antibody versus FITC Anti-CD45 antibody (Right Panel). The fluorescence histograms showing CD45 expression (or Ig Isotype control staining) were derived from gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.

Product Details
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BD Horizon™
Ptprc; LCA; Leukocyte common antigen; T200; Ly-5; Lyt-4
Mouse (QC Testing)
Rat LOU, also known as Louvain, LOU/C, LOU/M IgG2b, κ
Mouse Thymus / Spleen
Flow cytometry (Routinely Tested)
0.2 mg/ml
19264
AB_2738857
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. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  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.
564590 Rev. 4
Antibody Details
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30-F11

The 30-F11 clone has been reported to react with all isoforms and both alloantigens of CD45, which is found on hematopoietic stem cells and all cells of hematopoietic origin, except erythrocytes. CD45 is a transmembrane glycoprotein which is expressed at high levels on the cell surface, and its presence distinguishes leukocytes from non-hematopoietic cells. CD45 is a member of the Protein Tyrosine Phosphatase (PTP) family, where the intracellular carboxy-terminal region contains two PTP catalytic domains, and the extracellular region is highly variable due to alternative splicing of exons 4, 5, and 6 (designated as A, B, and C, respectively).  CD45 isoforms play complex roles in T-cell and B-cell antigen receptor signal transduction and the CD45 isoforms detected in the mouse are cell type-, maturation-, and activation state-specific.

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.

564590 Rev. 4
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.
altImg
BB515
Blue 488 nm
490 nm
515 nm
564590 Rev.4
Citations & References
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Development References (5)

  1. Johnson P, Maiti A, Ng DHW. CD45: A family of leukocyte-specific cell surface glycoproteins. In: Herzenberg LA, Weir DM, Herzenberg LA, Blackwell C , ed. Weir's Handbook of Experimental Immunology, Vol 2. Cambridge: Blackwell Science; 1997:62.1-62.16.
  2. Lagasse E, Connors H, Al-Dhalimy M, et al. Purified hematopoietic stem cells can differentiate into hepatocytes in vivo. Nat Med. 2000; 6(11):1212-1213. (Biology). View Reference
  3. Ledbetter JA, Herzenberg LA. Xenogeneic monoclonal antibodies to mouse lymphoid differentiation antigens. Immunol Rev. 1979; 47:63-90. (Immunogen: Cytotoxicity, Immunoprecipitation). View Reference
  4. Simon DI, Dhen Z, Seifert P, Edelman ER, Ballantyne CM, Rogers C. Decreased neointimal formation in Mac-1(-/-) mice reveals a role for inflammation in vascular repair after angioplasty. J Clin Pathol. 2000; 105(3):293-300. (Clone-specific: Immunohistochemistry). View Reference
  5. Thomas ML. The leukocyte common antigen family. Annu Rev Immunol. 1989; 7:339-369. (Clone-specific: Immunohistochemistry). View Reference
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564590 Rev. 4

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For Research Use Only. Not for use in diagnostic or therapeutic procedures.