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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.
- Human donor specific background has been observed in relation to the presence of anti-polyethylene glycol (PEG) antibodies, developed as a result of certain vaccines containing PEG, including some COVID-19 vaccines. We recommend use of BD Horizon Brilliant™ Stain Buffer in your experiments to help mitigate potential background. For more information visit https://www.bdbiosciences.com/en-us/support/product-notices.
Companion Products
The L203.rMab is a recombinant monoclonal antibody that was derived from L203 hybridoma cells. The L203.rMab specifically recognizes a monomorphic epitope on the extracellular region of human HLA-DR antigens which are human Major Histocompatibility Complex (MHC) Class II antigens. HLA-DR antigens are heterodimers comprised of two different type I transmembrane glycoproteins that are noncovalently-associated. The ~34 kDa HLA-DR alpha (HLA-DRα) chain is encoded by HLA-DRA whereas the ~28 kDa HLA-DR beta (HLA-DRβ) chains are encoded by one of the 4 different HLA-DRB loci (HLA-DRB1,3,4,5) that are located within the Human Leukocyte Antigen (HLA) Complex of chromosome 6. Each chain is comprised of an extracellular region with an IgSF domain, followed by a transmembrane sequence and a short cytoplasmic tail. The L203.rMab antibody recognizes a common determinant that is dependent on the association of HLA-DR alpha and beta chains. HLA-DR is variably expressed on B cells, activated T cells and NK cells, monocytes, macrophages, dendritic cells (DC), Langerhans cells, thymic epithelial cells, and tumor cell lines including B cell lines, myelomas, and some myeloid leukemias. HLA-DR functions in the presentation of peptide antigens to CD4+ T lymphocytes in the generation and regulation of adaptive immune responses. HLA-DR expressed on thymic stromal cells plays a key role in the positive and negative selection of CD4+ T cells during thymopoiesis. Certain HLA-DR alleles, polymorphisms or aberrant expression patterns are associated with susceptibility to diseases including autoimmunity and cancer. L203 antibody binding is reportedly blocked by the L243 monoclonal antibody suggesting that the two antibodies recognize crossreactive or spatially related determinants on HLA-DR.
Development References (4)
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Beck B, Dörfel D, Lichtenegger FS, et al. Effects of TLR agonists on maturation and function of 3-day dendritic cells from AML patients in complete remission.. J Transl Med. 2011; 9:151. (Clone-specific: Flow cytometry). View Reference
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Finn OJ, Levy R. Multiple HLA-DR antigens: detection with monoclonal antibodies and translation in vitro.. Proc Natl Acad Sci USA. 1982; 79(8):2658-62. (Clone-specific: Immunoprecipitation). View Reference
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Lampson LA, Levy R. Two populations of Ia-like molecules on a human B cell line.. J Immunol. 1980; 125(1):293-9. (Immunogen: Blocking, Immunoprecipitation, Radioimmunoassay). View Reference
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Suni MA, Picker LJ, Maino VC. Detection of antigen-specific T cell cytokine expression in whole blood by flow cytometry.. J Immunol Methods. 1998; 212(1):89-98. (Clone-specific: Blocking, Functional assay, Inhibition). 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.