Preparation and Storage
推奨アッセイ手順
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
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- 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.
- 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.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
関連製品
The B3B4 monoclonal antibody specifically binds to CD23, the low affinity IgE Fc receptor (FcεRII) expressed on mature resting conventional B lymphocytes, but not on B-1 cells (CD5+ B cells) or T lymphocytes. It does not react with high-affinity IgE receptors, as demonstrated on mouse mast cell lines. The regulation of CD23 surface expression on activated B cells appears to be complex, depending upon the mode of activation and the presence of cytokines. IgE synthesis is negatively regulated by CD23, and CD23 expression is upregulated on splenocytes in the presence of IgE. CD23 is also upregulated on follicular dendritic cells in the lymph nodes of immunized mice, and a subset of splenic dendritic cells expresses CD23. The B3B4 antibody abrogates antigen-specific IgE-dependent modulation of immune responses in normal mice. This monoclonal antibody also blocks IgE binding and eosinophil infiltration in the lung of immunized mice. Different in vivo results have been obtained when using the intact B3B4 antibody or the F(ab')2 fragments. B3B4 mAb does not cross-react with rat or human IgE Fc Receptor.
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.
Development References (14)
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Conrad DH, Waldschmidt TJ, Lee WT, et al. Effect of B cell stimulatory factor-1 (interleukin 4) on Fc epsilon and Fc gamma receptor expression on murine B lymphocytes and B cell lines. J Immunol. 1987; 139(7):2290-2296. (Clone-specific: Flow cytometry, Functional assay, Immunoaffinity chromatography, Immunoprecipitation, Radioimmunoassay). View Reference
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Coyle AJ, Wagner K, Bertrand C, Tsuyuki S, Bews J, Heusser C. Central role of immunoglobulin (Ig) E in the induction of lung eosinophil infiltration and T helper 2 cell cytokine production: inhibition by a non-anaphylactogenic anti-IgE antibody. J Exp Med. 1996; 183(4):1303-1310. (Clone-specific: Blocking). View Reference
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Dasic G, Juillard P, Graber P, et al. Critical role of CD23 in allergen-induced bronchoconstriction in a murine model of allergic asthma. Eur J Immunol. 1999; 29(9):2957-2967. (Clone-specific: Blocking, In vivo exacerbation). View Reference
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Kisselgof AB, Oettgen HC. The expression of murine B cell CD23, in vivo, is regulated by its ligand, IgE. Int Immunol. 1998; 10(9):1377-1384. (Clone-specific: Flow cytometry). View Reference
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Maeda K, Burton GF, Padgett DA, et al. Murine follicular dendritic cells and low affinity Fc receptors for IgE (Fc epsilon RII). J Immunol. 1992; 148(8):2340-2347. (Clone-specific: Electron microscopy, Immunohistochemistry). View Reference
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Oshiba A, Hamelmann E, Haczku A, et al. Modulation of antigen-induced B and T cell responses by antigen-specific IgE antibodies. J Immunol. 1997; 159(8):4056-4063. (Clone-specific: Flow cytometry). View Reference
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Pulendran B, Lingappa J, Kennedy MK, et al. Developmental pathways of dendritic cells in vivo: distinct function, phenotype, and localization of dendritic cell subsets in FLT3 ligand-treated mice. J Immunol. 1997; 159(5):2222-2231. (Clone-specific: Flow cytometry). View Reference
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Rabin E, Cong YZ, Wortis HH. Loss of CD23 is a consequence of B-cell activation. Implications for the analysis of B-cell lineages. Ann N Y Acad Sci. 1992; 651:130-142. (Biology: Flow cytometry). View Reference
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Rao M, Lee WT, Conrad DH. Characterization of a monoclonal antibody directed against the murine B lymphocyte receptor for IgE. J Immunol. 1987; 138(6):1845-1851. (Immunogen: Blocking, Immunoprecipitation, Inhibition, Radioimmunoassay). View Reference
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Stief A, Texido G, Sansig G, et al. Mice deficient in CD23 reveal its modulatory role in IgE production but no role in T and B cell development. J Immunol. 1994; 152(7):3378-3390. (Clone-specific: Flow cytometry, Immunohistochemistry). View Reference
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Waldschmidt T, Snapp K, Foy T, Tygrett L, Carpenter C. B-cell subsets defined by the Fc epsilon R. Ann N Y Acad Sci. 1992; 651:84-98. (Biology). View Reference
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Waldschmidt TJ, Conrad DH, Lynch RG. Expression of B cell surface receptors. II. IL-4 can accelerate the developmental expression of the murine B cell IgE Fc receptor. J Immunol. 1989; 143(9):2820-2827. (Clone-specific: Flow cytometry, Immunoaffinity chromatography). View Reference
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Waldschmidt TJ, Conrad DH, Lynch RG. The expression of B cell surface receptors. I. The ontogeny and distribution of the murine B cell IgE Fc receptor. J Immunol. 1988; 140(7):2148-2154. (Clone-specific: Flow cytometry). View Reference
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Yu P, Kosco-Vilbois M, Richards M, Kohler G, Lamers MC. Negative feedback regulation of IgE synthesis by murine CD23. Nature. 1994; 369(6483):753-756. (Biology). View Reference
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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.
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