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Flow cytometric analysis of Human CD274 expression by CD274-transfected Jurkat cells - Staining comparisons between BD Horizon™ BB515- and FITC-conjugated antibodies. Human MIT76 cells (CD274-transfected Jurkat cells) were stained with either BD Horizon BB515 Mouse IgG1, κ Isotype Control (Cat. No. 564416; dashed line histogram) or BD Horizon BB515 Mouse Anti-Human CD274 antibody (Cat. No. 564554; bold solid line histogram). Alternatively, cells were stained with FITC Anti-Human CD274 antibody (Cat. No. 558065; thin solid line histogram). Overlaid histograms are shown to facilitate staining comparisons between: BB515 Anti-CD274 antibody versus its Ig Isotype Control (Left Panel), and BB515 Anti-CD274 antibody versus FITC Anti-CD274 antibody (Right Panel). The fluorescence histograms showing CD274 expression (or Ig Isotype control staining) were derived from gated events with the forward and side light-scatter characteristics of viable cells. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.
BD Horizon™ BB515 Mouse Anti-Human CD274
Regulatory Status Legend
Any use of products other than the permitted use without the express written authorization of Becton, Dickinson and Company is strictly prohibited.
Preparation And Storage
Recommended Assay Procedures
BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® 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 BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
For optimal and reproducible results, BD Horizon Brilliant™ Stain Buffer should be used anytime BD Horizon Brilliant™ dyes are used in a multicolor flow cytometry panel. Fluorescent dye interactions may cause staining artifacts which may affect data interpretation. The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. When BD Horizon Brilliant Stain Buffer is used in in the multicolor panel, it should also be used in the corresponding compensation controls for all dyes to achieve the most accurate compensation. For the most accurate compensation, compensation controls created with either cells or beads should be exposed to BD Horizon Brilliant Stain Buffer for the same length of time as the corresponding multicolor panel. 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
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- 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).
- An isotype control should be used at the same concentration as the antibody of interest.
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- 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.
- Alexa Fluor™ is a trademark of Life Technologies Corporation.
Companion Products
The MIH1 monoclonal antibody specifically binds to CD274, which is also known as, B7 homolog 1 (B7-H1), Programmed cell death 1 ligand 1 (PDCD1 ligand, PDCD1L1, PDCD1LG1), or Programmed death ligand 1 (PD-L1, PDL1). CD274 and PD-L2 (CD273) are type I transmembrane glycoproteins that belong to the B7 family and serve as ligands for CD279 (Program Death 1/PD-1). CD274 is expressed on antigen-presenting cells including activated monocytes/macrophages and dendritic cells, as well as, activated T cells, and keratinocytes. CD274 is also expressed on placental trophoblasts, myocardial endothelium, cortical thymic epithelial cells, and on most carcinomas. CD274 plays an important role in regulating T cell responses. The MIH1 antibody blocks CD279 binding to CD274 and can enhance the proliferation and cytokine production of activated T cells.
Development References (7)
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Brown JA, Dorfman DM, Ma FR, et al. Blockade of programmed death-1 ligand on dendritic cells enhances T cell activation and cytokine production. J Immunol. 2003; 170:1257-1266. (Biology). View Reference
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Carreno BM, Bennett F, Chau TA, et al. CTLA-4 (CD152) can inhibit T cell activation by two different mechanisms depending on its level of cell surface expression.. J Immunol. 2000; 165(3):1352-6. (Biology). View Reference
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Carter L, Fouser LA, Jussif J, et al. PD-1:PD-L inhibitory pathway affects both CD4(+) and CD8(+) T cells and is overcome by IL-2. Eur J Immunol. 2002; 32:634-643. (Biology). View Reference
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Freeman GJ, Long AJ, Iwai Y, et al. Engagement of PD-1 immunoinhibitory receptor by a novel B7 family member leads to negative regulation of lymphocyte activation. J Exp Med. 2000; 192:1027-1034. (Biology). View Reference
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Latchman Y, Wood CR, Chernova T, et al. PD-L2 is a second ligand for PD-1 and inhibits T cell activation. Nat Immunol. 2001; 2(3):261-268. (Biology). View Reference
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Youngnak P, Kozono Y, Kozono H, et al. Differential binding properties of B7-H1 and B7-DC to programmed death-1. Biochem Biophys Res Commun. 2003; 307(3):672-677. (Immunogen: Flow cytometry). View Reference
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Youngnak-Piboonratanakit P, Tsushima F, Otsuki N, et al. The expression of B7-H1 on keratinocytes in chronic inflammatory mucocutaneous disease and its regulatory role. Immunol Lett. 2004; 94(3):215-222. (Clone-specific: Blocking, (Co)-stimulation, Flow cytometry, Fluorescence microscopy, Functional assay, Immunofluorescence, Immunohistochemistry). View Reference
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.