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BB515 Rat Anti-Mouse CD278
BB515 Rat Anti-Mouse CD278
Two-color flow cytometric analysis of CD278 (ICOS) expression on activated mouse splenocytes. Mouse splenic leucocytes were either unstimulated (Top Panels) or activated (3 days; Bottom Panels) by plate-bound Purified NA/LE Hamster Anti-Mouse CD3e antibody (Cat. No. 553057). The cells were harvested and preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). Both sets of cells were stained with APC Rat Anti-Mouse CD8a antibody (Cat. No. 553035/561093) and either BD Horizon™ BB515 Rat IgG2b, κ Isotype Control (Cat. No. 564421; Left Panels) or BD Horizon BB515 Rat Anti-Mouse CD278 antibody (Cat. No. 564592; Right Panels). Two-color flow cytometric contour plots showing the correlated expression patterns of CD278 (or Ig Isotype control staining) versus CD8a were derived for gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Two-color flow cytometric analysis of CD278 (ICOS) expression on activated mouse splenocytes. Mouse splenic leucocytes were either unstimulated (Top Panels) or activated (3 days; Bottom Panels) by plate-bound Purified NA/LE Hamster Anti-Mouse CD3e antibody (Cat. No. 553057). The cells were harvested and preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). Both sets of cells were stained with APC Rat Anti-Mouse CD8a antibody (Cat. No. 553035/561093) and either BD Horizon™ BB515 Rat IgG2b, κ Isotype Control (Cat. No. 564421; Left Panels) or BD Horizon BB515 Rat Anti-Mouse CD278 antibody (Cat. No. 564592; Right Panels). Two-color flow cytometric contour plots showing the correlated expression patterns of CD278 (or Ig Isotype control staining) versus CD8a were derived for gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Product Details
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BD Horizon™
ICOS; Ailim; CCLP; CRP-1; Ly115; Inducible T-cell costimulator
Mouse (QC Testing)
Rat IgG2b, κ
Mouse Icos cDNA and ICOS hexahistidine fusion protein
Flow cytometry (Routinely Tested)
0.2 mg/ml
AB_2738858
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. 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.
566214 Rev. 2
Antibody Details
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7E.17G9

The 7E.17G9 monoclonal antibody specifically binds to CD278, the Inducible Costimulatory molecule (ICOS), a 47-57 kDa homodimeric glycoprotein of the CD28 family of costimulatory molecules. ICOS is expressed on subpopulations of CD4-CD8- and CD4+CD8- (but not CD4-CD8+ or CD4+CD8+) thymocytes, on some T-cell lines, and on small numbers of peripheral leukocytes. It is upregulated on T lymphocytes following activation via the T-cell receptor. The T-cell activation marker H4 is the same molecule as  ICOS. ICOS is a costimulatory receptor, and its ligand on antigen-presenting cells has been called B7RP-1, GL50, B7h, B7-H2, or LICOS. There is considerable evidence that the interaction of ICOS with its ligand is involved in the regulation of many, but not all, T-cell-mediated immune responses.

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.

566214 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
566214 Rev.2
Citations & References
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Development References (8)

  1. Buonfiglio D, Bragardo M, Redoglia V, et al. The T cell activation molecule H4 and the CD28-like molecule ICOS are identical. Eur J Immunol. 2000; 30(12):3463-3467. (Biology). View Reference
  2. Chambers CA. The expanding world of co-stimulation: the two-signal model revisited. Trends Immunol. 2001; 22(4):217-223. (Biology). View Reference
  3. Dong C, Temann UA, Flavell RA. Cutting edge: critical role of inducible costimulator in germinal center reactions. J Immunol. 2001; 166(6):3659-3662. (Biology). View Reference
  4. Mages HW, Hutloff A, Heuck C, et al. Molecular cloning and characterization of murine ICOS and identification of B7h as ICOS ligand. Eur J Immunol. 2000; 30(4):1040-1047. (Biology). View Reference
  5. McAdam AJ, Chang TT, Lumelsky AE, et al. Mouse inducible costimulatory molecule (ICOS) expression is enhanced by CD28 costimulation and regulates differentiation of CD4+ T cells.. J Immunol. 2000; 165(9):5035-40. (Immunogen: ELISA, Flow cytometry). View Reference
  6. Schwartz RH. Immunology. It takes more than two to tango. Nature. 2001; 409(6816):31-32. (Biology). View Reference
  7. Sperling AI, Bluestone JA. ICOS costimulation: It's not just for TH2 cells anymore. Nat Immunol. 2001; 2(7):573-574. (Biology). View Reference
  8. Wallin JJ, Liang L, Bakardjiev A, Sha WC. Enhancement of CD8+ T cell responses by ICOS/B7h costimulation. J Immunol. 2001; 167(1):132-139. (Biology). View Reference
View All (8) View Less
566214 Rev. 2

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