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 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® CompBeads are appropriate for your specific cellular application.
For optimal and reproducible results, BD Horizon Brilliant™ Stain Buffer should be used anytime BD Horizon Brilliant™ dyes are used in a multicolor flow cytometry panel. Fluorescent dye interactions may cause staining artifacts which may affect data interpretation. The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. When BD Horizon Brilliant Stain Buffer is used in in the multicolor panel, it should also be used in the corresponding compensation controls for all dyes to achieve the most accurate compensation. For the most accurate compensation, compensation controls created with either cells or beads should be exposed to BD Horizon Brilliant Stain Buffer for the same length of time as the corresponding multicolor panel. 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 two 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
- 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).
- 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.
- This product may be covered by US Patent No. 7,388,080.
- 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.
- 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.
- Alexa Fluor™ is a trademark of Life Technologies Corporation.
Companion Products
The 11G7 monoclonal antibody specifically binds to human CD282, which is also known as Toll-like receptor 2 (TLR2). CD282 is expressed on monocytes, granulocytes, and dendritic cells. Toll-like receptors (TLRs) play a critical role in antimicrobial resistance. Moreover, TLRs have been shown to activate a number of signal transduction pathways which lead to the induction of genes involved in host defense. TLRs are type-1 transmembrane receptors characterized by the presence of extracellular leucine-rich repeat and intracellular Toll/IL-1 receptor domains. At least 12 mammalian TLRs have been identified, each recognizing a distinct bacterial or viral pathogen-associated molecular pattern, termed PAMP. Peptidoglycan from Gram-positive bacteria, lipoproteins and lipopeptides from several bacteria, glycophosphatidylinositol, lipoarabinomannan, porins, and zymosan from yeast have been reported to be the ligands for TLR2.
It has been reported that mAb 11G7 inhibits the production of inflammatory cytokines via certain TLR2 ligands including TLR2/TLR1 ligands, lipoarabinomannan and PAM3CSK4. However, 11G7 antibody does not inhibit the production of inflammatory cytokines with zymosan, a TLR2/TLR6 ligand. Please note that this application has not been tested at BD Biosciences Pharmingen.
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 (10)
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Akira, S.. Toll-Like Receptor signaling. Immunology. 2004; 4:499-511. (Biology).
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Buhring HJ, Saalmuller A, Muller C, van Agthoven AJ, Busch FW. The monoclonal antibody 11G7 recognizes a novel differentiation antigen expressed on hemopoietic precursor cells. Hybridoma. 1991; 10(1):77-88. (Biology). View Reference
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Kurt-Jones EA, Mandell L, Whitney C, et al. Role of Toll-like receptor 2 (TLR2) in neutrophil activation: GM-CSF enhances TLR2 expression and TLR2-mediated interleukin 8 responses in neutrophils. Blood. 2002; 100(5):1860-1868. (Biology). View Reference
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Lien E, Sellati TJ, Yoshimura A, et al. Toll-like receptor 2 functions as a pattern recognition receptor for diverse bacterial products. J Biol Chem. 1999; 274(47):33419-33425. (Biology). View Reference
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Medzhitov R. Toll-like receptors and innate immunity. Nat Rev Immunol. 2001; 1(2):135-145. (Biology). View Reference
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Muzio, M.. Toll-Like Receptor family and signaling pathway. Biochem J. 2000; 28:563-566. (Biology).
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Nilsen N, Nonstad U, Khan N, et al. Lipopolysaccharide and double-stranded RNA up-regulate toll-like receptor 2 independently of myeloid differentiation factor 88.. J Biol Chem. 2004; 279(38):39727-35. (Biology). View Reference
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Paterson, H.. Injury Primes the innate immune system for enhanced Toll-Like Receptor reactivity. J Immunol. 2003; 171:1473-1483. (Biology).
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Sandor F, Latz E, Re F, et al. Importance of extra- and intracellular domains of TLR1 and TLR2 in NFκB signaling. J Cell Biol. 2003; 162(6):1099-1110. (Clone-specific: Blocking, Inhibition). View Reference
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Zhou S, Cerny AM, Bowen G, et al. Discovery of a novel TLR2 signaling inhibitor with anti-viral activity.. Antiviral Res. 2010; 87(3):295-306. (Biology). View Reference
Please refer to Support Documents for Quality Certificates
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.
