BUV563 Rat Anti-Mouse Ly-49D
Clone 4E5 (RUO)
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- Alternative Name Ly49d; Ly-49d; Klra4; Chok
- Concentration 0.2 mg/ml
- Isotype Rat F344, also known as Fischer, CDF IgG2a, κ
- Reactivity Mouse (Tested in Development)
Flow cytometry (Qualified)
- Immunogen Not Reported
- Storage Buffer Aqueous buffered solution containing ≤0.09% sodium azide.
- Regulatory Status RUO
Regulatory Status Legend
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.
BUV563 is a tandem fluorochrome that combines BD Horizon BUV395 and an acceptor dye with an EM Max at 563 nm. Due to the excitation of the acceptor dye by other laser lines, there may be spillover into channels detecting PE (eg, 575/26-nm filter) and PE-CF594 (eg, 610/20-nm filter). BUV563 has been exclusively developed by BD Biosciences for instruments equipped with a 355-nm UV laser.
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Preparation and Storage
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.
The antibody was conjugated with BD Horizon BUV563 under optimal conditions that minimize unconjugated dye and antibody.
- 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.
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).