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BB700 Mouse Anti-Human CD95
BB700 Mouse Anti-Human CD95
Multiparameter flow cytometric analysis of CD95 expression on human peripheral blood leucocyte populations. Human whole blood was stained with either BD Horizon™ BB700 Mouse IgG1, κ Isotype Control (Cat. No. 566404; Left Plot) or BD Horizon BB700 Mouse Anti-Human CD95 antibody (Cat. No. 566542/566543; Right Plot). A two-parameter flow cytometric dot plot showing the correlated expression of CD95 (or Ig Isotype control staining) versus side-light scatter (SSC-A) signals was derived from gated events with the forward and side-light scatter characteristics of intact leucocyte populations. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.
Multiparameter flow cytometric analysis of CD95 expression on human peripheral blood leucocyte populations. Human whole blood was stained with either BD Horizon™ BB700 Mouse IgG1, κ Isotype Control (Cat. No. 566404; Left Plot) or BD Horizon BB700 Mouse Anti-Human CD95 antibody (Cat. No. 566542/566543; Right Plot). A two-parameter flow cytometric dot plot showing the correlated expression of CD95 (or Ig Isotype control staining) versus side-light scatter (SSC-A) signals was derived from gated events with the forward and side-light scatter characteristics of intact leucocyte populations. Flow cytometric analysis was performed using a BD LSRFortessa™ Cell Analyzer System.
Product Details
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BD Horizon™
APO-1; FAS; TNFRSF6; APT1; ALPS1A; FAS1; FASTM; FASLG receptor
Human (QC Testing), Rhesus, Cynomolgus, Baboon (Tested in Development)
Mouse C3H, also known as C3H/He, C3H/Bi IgG1, κ
Human CD95-transfected L Cells
Flow cytometry (Routinely Tested)
5 µl
VI C-64
355
AB_2869780
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™ BB700 under optimum conditions, and unconjugated antibody and free BD Horizon BB700 were removed.

Recommended Assay Procedures

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 for the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).

When setting up compensation, it is recommended to compare spillover values obtained from cells and BD™ CompBeads to ensure that beads will provide sufficiently accurate spillover values.

For optimal results, it is recommended to perform two washes after staining with antibodies. Cells may be prepared, stained with antibodies and washed twice with wash buffer per established protocols for immunofluorescent 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. The manufacture, use, sale, offer for sale, or import of this product is subject to one or more patents or pending applications. This product, and only in the amount purchased by buyer, may be used solely for buyer’s own internal research, in a manner consistent with the accompanying product literature. No other right to use, sell or otherwise transfer (a) this product, or (b) its components is hereby granted expressly, by implication or by estoppel. Diagnostic uses require a separate license.
  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. BD Horizon Brilliant Blue 700 is covered by one or more of the following US patents: 8,455,613 and 8,575,303.
  8. Cy is a trademark of GE Healthcare.
  9. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  10. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
566542 Rev. 1
Antibody Details
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DX2

The DX2 monoclonal antibody specifically binds to the human Fas antigen (also called APO-1). This 45 kDa type I transmembrane glycoprotein was designated as CD95 at the Fifth HLDA Workshop. Fas is a member of the TNF-receptor superfamily and is also known as Tumor necrosis factor receptor superfamily member 6 (TNFRSF6). It is differentially expressed on a variety of normal and neoplastic cells. These include some undifferentiated thymocytes, and activated T and B lymphocytes, natural killer (NK) cells, monocytes, neutrophils, fibroblasts, and cell lines. CD95 is preferentially expressed on CD45RO-positive memory T lymphocytes and γ/δ T lymphocytes. The Fas/CD95 antigen is a polypeptide that plays a role in the programmed sequence of events leading to cell death, termed apoptosis. Crosslinking CD95 with DX2 antibody delivers an apoptotic signal indicating that DX2 recognizes a functional epitope of the CD95 antigen.

The antibody was conjugated to BD Horizon BB700, which is part of the BD Horizon Brilliant™ Blue family of dyes.   It is a polymer-based tandem dye developed exclusively by BD Biosciences.  With an excitation max of 485 nm and an emission max of 693 nm, BD Horizon BB700 can be excited by the 488 nm laser and detected in a standard PerCP-Cy™5.5 set (eg, 695/40-nm filter). This dye provides a much brighter alternative to PerCP-Cy5.5 with less cross laser excitation off the 405 nm and 355 nm lasers.

566542 Rev. 1
Format Details
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BB700
The BD Horizon Brilliant™ Blue 700 (BB700) Dye is part of the BD Horizon Brilliant™ Blue family of dyes. This tandem fluorochrome is comprised of a polymer-technology dye donor with an excitation maximum (Ex Max) of 476-nm and an acceptor dye with an emission maximum (Em Max) at 695-nm. Driven by BD innovation, BB700 is designed to be excited by the blue laser (488-nm) and detected using an optical filter centered near 695-nm (e.g., a 695/20-nm bandpass filter). The donor dye can be excited by the Violet (405 nm) laser and the acceptor dye can be excited by the red (627–640 nm) laser resulting in cross-laser excitation and fluorescence spillover. BB700 Reagents are significantly brighter than equivalent PerCP or PerCP-Cy5.5 reagents and are less sensitive to photobleaching. In addition, BB700 shows much less excitation by the violet (407-nm) laser resulting in less spillover. BB700 has minimal yellow green (562-nm) excitation and is ideal for instruments with both blue (488-nm) and yellow green (562-nm) lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BB700
Blue 488 nm
476 nm
695 nm
566542 Rev.1
Citations & References
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Development References (6)

  1. Cifone MG, De Maria R, Roncaioli P, et al. Apoptotic signaling through CD95 (Fas/Apo-1) activates an acidic sphingomyelinase. J Exp Med. 1994; 180(4):1547-1552. (Immunogen: Apoptosis, Functional assay). View Reference
  2. Itoh N, Yonehara S, Ishii A, et al. The polypeptide encoded by the cDNA for human cell surface antigen Fas can mediate apoptosis. Cell. 1991; 66(2):233-243. (Biology). View Reference
  3. 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.
  4. Lanier LL, Chang C, Phillips JH. Human NKR-P1A. A disulfide-linked homodimer of the C-type lectin superfamily expressed by a subset of NK and T lymphocytes. J Immunol. 1994; 153(6):2417-2428. (Clone-specific: Flow cytometry). View Reference
  5. Robertson MJ, Ritz R. Cluster report: CD95. In: 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; 1995:1142-1143.
  6. Yonehara S, Nakamachi Y. CD95 Workshop Panel Report. In: 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.; 1997:804-814.
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566542 Rev. 1

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