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BB700 Mouse Anti-Human CD3
BB700 Mouse Anti-Human CD3

Flow cytometric analysis of CD3 expression on Rhesus monkey peripheral blood lymphocytes. Rhesus whole blood was stained with either BD Horizon™ BB700 Mouse IgG1, κ Isotype Control (Cat. No. 566404; dashed line histogram) or BD Horizon BB700 Mouse Anti-Human CD3 antibody (Cat. No. 566517/566518; solid line histogram) at 1 µg/test. Erythrocytes were lysed with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899). The normalized fluorescence histograms showing CD3 expression or Ig Isotype control staining were derived from gated events with the forward and side light-scatter characteristics of viable lymphocytes. Flow cytometric analysis was performed using a BD FACSCelesta™ Flow Cytometer System. Data shown on this Technical Data Sheet are not lot specific.

Flow cytometric analysis of CD3 expression on Rhesus monkey peripheral blood lymphocytes. Rhesus whole blood was stained with either BD Horizon™ BB700 Mouse IgG1, κ Isotype Control (Cat. No. 566404; dashed line histogram) or BD Horizon BB700 Mouse Anti-Human CD3 antibody (Cat. No. 566517/566518; solid line histogram) at 1 µg/test. Erythrocytes were lysed with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899). The normalized fluorescence histograms showing CD3 expression or Ig Isotype control staining were derived from gated events with the forward and side light-scatter characteristics of viable lymphocytes. Flow cytometric analysis was performed using a BD FACSCelesta™ Flow Cytometer System. Data shown on this Technical Data Sheet are not lot specific.

Product Details
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BD Horizon™
CD3E; CD3-epsilon; T3E; TCRE
Rhesus, Cynomolgus, Baboon (QC Testing), Human (Tested in Development)
Mouse BALB/c IgG1, λ
Purified Human CD3ε Protein
Flow cytometry (Routinely Tested)
0.2 mg/ml
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. 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. 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. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
566518 Rev. 1
Antibody Details
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SP34-2

Clone SP34-2 is a mouse IgG1 isotype monoclonal antibody, descendant of SP34 (mouse IgG3), with the same specificity and reactivity pattern as the parent clone. It cross-reacts with a major subset of peripheral blood lymphocytes, but not monocytes or granulocytes, of baboon, and rhesus, cynomolgus, and pigtail macaque monkeys. The distribution on lymphocytes is similar to that observed with normal human donor lymphocytes with the majority of CD3-positive cells being negative when dual stained with antibodies to B or NK cells markers. SP34-2 is also capable of inducing cell proliferation on both human and non-human primate PBMC.

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.

566518 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
566518 Rev.1
Citations & References
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Development References (7)

  1. Bleavins MR, Brott DA, Alvey JD, de la Iglesia FA. Flow cytometric characterization of lymphocyte subpopulations in the cynomolgus monkey (Macaca fascicularis). Vet Immunol Immunopathol. 1993; 37(1):1-13. (Biology). View Reference
  2. Blumberg RS, Ley S, Sancho J, et al. Structure of the T-cell antigen receptor: evidence for two CD3 epsilon subunits in the T-cell receptor-CD3 complex. Proc Natl Acad Sci U S A. 1990; 87(18):7220-7224. (Clone-specific: Immunoprecipitation, Western blot). View Reference
  3. Conrad ML, Davis WC, Koop BF. TCR and CD3 antibody cross-reactivity in 44 species. Cytometry A. 2007; 71(11):925-933. (Clone-specific: Flow cytometry). View Reference
  4. Engel P, Wagner N, Tedder TF. CD86 Workshop Report. 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; 1995:703-705.
  5. Jacobsen CN, Aasted B, Broe MK, Petersen JL. Reactivities of 20 anti-human monoclonal antibodies with leucocytes from ten different animal species. Vet Immunol Immunopathol. 1993; 39(4):461-466. (Biology). View Reference
  6. Lin G-X, Yang X, Hollemweguer E, et al. Cross-reactivity of CD antibodies in eight animal species. In: Mason D. David Mason .. et al., ed. Leucocyte typing VII : white cell differentiation antigens : proceedings of the Seventh International Workshop and Conference held in Harrogate, United Kingdom. Oxford: Oxford University Press; 2002:519-523.
  7. Pessano S, Oettgen H, Bhan AK, Terhorst C. The T3/T cell receptor complex: antigenic distinction between the two 20-kd T3 (T3-delta and T3-epsilon) subunits. EMBO J. 1985; 4(2):337-344. (Immunogen: Activation, Dot Blot, Functional assay, Immunoprecipitation, Western blot). View Reference
View All (7) View Less
566518 Rev. 1

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