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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.
Product Notices
- When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
- Researchers should determine the optimal concentration of this reagent for their individual applications.
- The production process underwent stringent testing and validation to assure that it generates a high-quality conjugate with consistent performance and specific binding activity. However, verification testing has not been performed on all conjugate lots.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
- 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.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
Companion Products
CD244 (also known as 2B4 Antigen) is a member of the CD2 subset of the immunoglobulin superfamily (CD2 IgSF). It is expressed on all natural killer (NK) cells, IL-2–activated NK cells (LAK), and a subset of T lymphocytes which mediate non-MHC–restricted cytotoxicity. The C9.1 antibody specifically recognizes CD244.1, the 2B4 alloantigen which is expressed by certain cells in most strains tested (A/J, AKR/N, BALB/c, CBA/N, C3H/He, DBA/1, DBA/2, NZB/N, SJL/J, 129/Svj), but not by cells from C57BL/6 or C58/J mice. The expression of the CD244.1 alloantigen on B lymphocytes that was originally described, has not been confirmed in later studies. The extracellular domain of CD244 is a ligand for another CD2 IgSF member, CD48.
Development References (7)
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Brown MH, Boles K, van der Merwe PA, Kumar V, Mathew PA, Barclay AN. 2B4, the natural killer and T cell immunoglobulin superfamily surface protein, is a ligand for CD48. J Exp Med. 1998; 188(11):2083-2090. (Biology). View Reference
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Davis SJ, van der Merwe PA. The structure and ligand interactions of CD2: implications for T-cell function. Immunol Today. 1996; 17(4):177-187. (Biology). View Reference
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Kubota K, Katoh H, Muguruma K, Koyama K. Characterization of a surface membrane molecule expressed by natural killer cells in most inbred mouse strains: monoclonal antibody C9.1 identifies an allelic form of the 2B4 antigen. Immunology. 1999; 96(3):491-497. (Immunogen: Cytotoxicity, Depletion, Flow cytometry, Fluorescence activated cell sorting, Immunoprecipitation). View Reference
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Kubota K. A killer cell protective antigen expressed by MHC-unrestricted killer hybridomas. Cell Immunol. 1997; 181(1):50-58. (Biology). View Reference
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Kumaresan PR, Huynh VT, Mathew PA. Polymorphism in the 2B4 gene of inbred mouse strains. Immunogenetics. 2000; 51(8-9):758-761. (Biology). View Reference
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Latchman Y, McKay PF, Reiser H. Identification of the 2B4 molecule as a counter-receptor for CD48. J Immunol. 1998; 161(11):5809-5812. (Biology). View Reference
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Mathew PA, Garni-Wagner BA, Land K, et al. Cloning and characterization of the 2B4 gene encoding a molecule associated with non-MHC-restricted killing mediated by activated natural killer cells and T cells. J Immunol. 1993; 151(10):5328-5337. (Biology). View Reference
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Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described
Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.