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Regulatory Status Legend
Any use of products other than the permitted use without the express written authorization of Becton, Dickinson and Company is strictly prohibited.
Preparation And Storage
Recommended Assay Procedures
For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes (including BD OptiBuild Brilliant reagents) 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).
Product Notices
- This antibody was developed for use in flow cytometry.
- 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.
- Researchers should determine the optimal concentration of this reagent for their individual applications.
- 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.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- 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.
Companion Products
The 4E5 monoclonal antibody specifically recognizes Ly-49D, which is expressed on subsets of natural killer (NK) cells in C57BL/6, C3H/He (at a very low frequency), and SJL, but not DBA/2, AKR, CBA/J, or BALB/c mice. Unlike Ly-49D antigen has not been betected on NK-1.1+ (or DX5+) T cells. In 129/J mice, the 4E5 antibody cross-reacts with Ly-49O, Ly-49R, and Ly-49V. The Ly-49 family of NK-cell receptors, members of the C-type lectin superfamily, are disulfide-linked type-II transmembrane protein homodimers with extracellular carbohydrate-recognition domains, which bind to MHC class I alloantigens. The Ly-49 family members are expressed independently, such that an individual NK or T cell may display more than one class of Ly-49 receptor homodimers. Ly-49D weakly binds to MHC class I antigens of the k halpotype, and Ly-49D+ IL-2-activated NK cells lyse target cells expressing H-2[a], H-2[b], H-2[d], H-2[k], H-2[p], H-2[q], and H-2[s] and the CHO (Chinese hamster ovary) cell line. Ly-49D+ cells mediate allogenic resistance to H-2d bone marrow transplantation. In vitro studies suggest that the Ly-49D receptor mediates activation of NK-cell cytolytic activity via tyrosine phosphorylation of their ITIMs (Immunoreceptor Tyrosine-based Inhibitory Motifs). Molecular differences between the Ly-49D stimulatory receptor and the inhibitory members of the Ly-49 family include the absence of an ITIM in Ly-49D, the lack of phosphorylation of Ly-49D in activated NK cells, and the association of a novel tyrosine-phosphorylated protein (pp16) with Ly-49D in activated NK cells. Ly-49O and Ly-49V are closely related to Ly-49A[B6] and, like Ly-49A, have ITIM domains. Ly-49O- and Ly-49V-transfected 293T (human kidney epithelial) cells bind tetramers of H-2D[b], D[b], D[k], and L[d]. In addition, the Ly-49V-transfected cells also bind K[b], K[d], and K[k]. Ly-49R is closely related to Ly-49D[B6] and is putative activating receptor due to its lack of an ITIM domain. Ly-49R-transfected 293T cells bind soluble tetramers of H-2D[b], D[d], D[k], and L[d].
The antibody was conjugated to BD Horizon™ BUV563 which is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This dye is a tandem fluorochrome of BD Horizon BUV395 which has an Ex Max of 348 nm and an acceptor dye. The tandem has an Em Max at 563 nm. BD Horizon BUV563 can be excited by the 355 nm ultraviolet laser. On instruments with a 561 nm Yellow-Green laser, the recommended bandpass filter is 585/15 nm with a 535 nm long pass to minimize laser light leakage. When BD Horizon BUV563 is used with an instrument that does not have a 561 nm laser, a 560/40 nm filter with a 535 nm long pass may be more optimal. Due to the excitation and emission characteristics of the acceptor dye, there may be spillover into the PE and PE-CF594 detectors. However, the spillover can be corrected through compensation as with any other dye combination.
Development References (13)
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George TC, Mason LH, Ortaldo JR, Kumar V, Bennett M. Positive recognition of MHC class I molecules by the Ly49D receptor of murine NK cells. J Immunol. 1999; 162(4):2035-2043. (Biology). View Reference
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Hanke T, Takizawa H, McMahon CW, et al. Direct assessment of MHC class I binding by seven Ly49 inhibitory NK cell receptors. Immunity. 1999; 11(1):67-77. (Biology). View Reference
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Idris AH, Smith HR, Mason LH, Ortaldo JR, Scalzo AA, Yokoyama WM. The natural killer gene complex genetic locus Chok encodes Ly-49D, a target recognition receptor that activates natural killing. Proc Natl Acad Sci U S A. 1999; 96(11):6330-6335. (Biology). View Reference
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Makrigiannis AP, Pau AT, Saleh A, Winkler-Pickett R, Ortaldo JR, Anderson SK. Class I MHC-binding characteristics of the 129/J Ly49 repertoire. J Immunol. 2001; 166(8):5034-5043. (Biology). View Reference
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Mason LH, Anderson SK, Yokoyama WM, Smith HR, Winkler-Pickett R, Ortaldo JR. The Ly-49D receptor activates murine natural killer cells. J Exp Med. 1996; 184(6):2119-2128. (Biology). View Reference
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Mason LH, Gosselin P, Anderson SK, Fogler WE, Ortaldo JR, McVicar DW. Differential tyrosine phosphorylation of inhibitory versus activating Ly-49 receptor proteins and their recruitment of SHP-1 phosphatase. J Immunol. 1997; 159(9):4187-4196. (Biology). View Reference
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Mason LH, Willette-Brown J, Anderson SK, et al. Characterization of an associated 16-kDa tyrosine phosphoprotein required for Ly-49D signal transduction. J Immunol. 1998; 160(9):4148-4152. (Biology). View Reference
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Ortaldo JR, Mason AT, Winkler-Pickett R, Raziuddin A, Murphy WJ, Mason LH. Ly-49 receptor expression and functional analysis in multiple mouse strains. J Leukoc Biol. 1999; 66(3):512-520. (Clone-specific). View Reference
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Ortaldo JR, Winkler-Pickett R, Mason AT, Mason LH. The Ly-49 family: regulation of cytotoxicity and cytokine production in murine CD3+ cells. J Immunol. 1998; 160(1):1158-1165. (Clone-specific). View Reference
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Raulet DH, Held W, Correa I, Dorfman JR, Wu MF, Corral L. Specificity, tolerance and developmental regulation of natural killer cells defined by expression of class I-specific Ly49 receptors. Immunol Rev. 1997; 155:41-52. (Biology). View Reference
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Raziuddin A, Longo DL, Mason L, Ortaldo JR, Bennett M, Murphy WJ. Differential effects of the rejection of bone marrow allografts by the depletion of activating versus inhibiting Ly-49 natural killer cell subsets. J Immunol. 1998; 160(1):87-94. (Biology). View Reference
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Takei F, Brennan J, Mager DL. The Ly-49 family: genes, proteins and recognition of class I MHC. Immunol Rev. 1997; 155:67-77. (Biology). View Reference
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Yokoyama WM, Seaman WE. The Ly-49 and NKR-P1 gene families encoding lectin-like receptors on natural killer cells: the NK gene complex. Annu Rev Immunol. 1993; 11:613-635. (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.