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 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.
- 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.
- 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.
Companion Products
The CW-1.2 monoclonal antibody specifically binds to CD199 which is also known as Chemokine (C-C motif) receptor 9 (CCR9) or C-C chemokine receptor type 9 (C-C CKR-9). CD199 is a ~42 kDa seven-transmembrane protein and member of the G protein-coupled receptor 1 family. CD199 binds to CCL25, which is likewise known as thymus-expressed chemokine (TECK) or small inducible chemokine 25 (Scya25). CCL25 is selectively and constitutively expressed in the thymic cortex and small intestinal epithelium. CD199 is highly expressed on CD4+CD8+ double-positive thymocytes, and mature naïve CD8+ T cells, but not naïve CD4+ T cells, in the peripheral lymphoid organs. CD199 is also expressed on small intestinal B cells, and on subsets of memory CD4+ T cells and CD8+ T cells, and dendritic cells. The CCR9/CCL25 pathway plays important roles in T cell development and gut-associated immune functions. It is especially involved in the recruitment of CD8αα+ intraepithelial lymphocytes (IELs) and the homing of other lymphocytes to the gut. Dysregulation of this pathway is associated with inflammatory responses, such as inflammatory bowel disease (IBD) and celiac disease.
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 (7)
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Drakes ML, Stiff PJ, Blanchard TG. Inverse relationship between dendritic cell CCR9 expression and maturation state. Immunology. 2009; 127(4):466-476. (Biology). View Reference
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Feng N, Jaimes MC, Lazarus NH, et al. Redundant role of chemokines CCL25/TECK and CCL28/MEC in IgA+ plasmablast recruitment to the intestinal lamina propria after rotavirus infection. J Immunol. 2006; 176(10):5749-5759. (Biology). View Reference
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Hadeiba H, Lahl K, Edalati A, et al. Plasmacytoid dendritic cells transport peripheral antigens to the thymus to promote central tolerance. Immunity. 2012; 36(3):438-450. (Biology). View Reference
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McGuire HM, Vogelzang A, Ma CS, et al. A subset of interleukin-21+ chemokine receptor CCR9+ T helper cells target accessory organs of the digestive system in autoimmunity. Immunity. 2011; 34(4):602-615. (Biology). View Reference
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Wendland M, Czeloth N, Mach N, et al. CCR9 is a homing receptor for plasmacytoid dendritic cells to the small intestine. Proc Natl Acad Sci U S A. 2007; 104(15):6347-6352. (Biology). View Reference
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Wurbel MA, Malissen B, Campbell JJ. Complex regulation of CCR9 at multiple discrete stages of T cell development. Eur J Immunol. 2006; 36(1):73-81. (Immunogen: Flow cytometry). View Reference
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Wurbel MA, Malissen M, Guy-Grand D, Malissen B, Campbell JJ. Impaired accumulation of antigen-specific CD8 lymphocytes in chemokine CCL25-deficient intestinal epithelium and lamina propria. J Immunol. 2007; 178(12):7598-7606. (Clone-specific: Flow cytometry). 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.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.