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 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 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).
Note: When using high concentrations of antibody, background binding of this dye to erythroid cell subsets (mature erythrocytes and precursors) has been observed. For researchers studying these cell populations, or in cases where light scatter gating does not adequately exclude these cells from the analysis, this background may be an important factor to consider when selecting reagents for panel(s).
Product Notices
- 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.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- BD Horizon Brilliant Ultraviolet 496 is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,227,187; 8,575,303; 8,354,239.
Companion Products
The HP-MA4 monoclonal antibody specifically recognizes several Killer Cell Immunoglobulin-like Receptors (KIRs) which are also known as CD158 molecules. HP-MA4 recognizes Killer cell immunoglobulin-like receptor 2DL1 (encoded by KIR2DL1; aka, CD158a and NKAT-1), Killer cell immunoglobulin-like receptor 2DS1 (KIR2DS1; CD158h), Killer cell immunoglobulin-like receptor 2DS3 (KIR2DS3; NKAT-7), and Killer cell immunoglobulin-like receptor 2DS5 (KIR2DS5; CD158g, NKAT-9) which are collectively known as KIR2DL1/S1/S3/S5 (CD158). These type I transmembrane glycoproteins are encoded by polymorphic genes and have 2 extracellular Ig-like domains (KIR2D, domains D1 and D2) followed by a transmembrane region and either long (L) or short (S) cytoplasmic domains. Various CD158 molecules are differentially expressed by CD56dim natural killer (NK) cells and some T cells and can regulate their cytotoxic effector functions. Although different KIR gene content varies amongst haplotypes for individuals, certain "framework" genes including KIR3DL3, KIR3DP1, KIR3DL4, and KIR3DL2, are found in all haplotypes. KIR2DL1 has a long cytoplasmic domain with two tyrosine-based inhibitory motifs (ITIM) that enables inhibitory signal transduction by ligand-bound KIR2DL1 leading to reduced cytotoxic effector cell activity. KIR2DS1, KIR2DS3, KIR2DS5 (KIR2DS1/S3/S5) proteins each have a short cytoplasmic tail with a positively charged amino acid in their transmembrane region which allows association with the DAP12 transmembrane protein. DAP12 acts as an activating signal transduction element through its immunoreceptor tyrosine-based activation motifs (ITAMs) in its cytoplasmic domain leading to upregulated cytotoxic effector cell function. Some MHC class I molecules can serve as ligands for CD158 molecules, with HLA-C ligands reported for KIR2DL1, KIR2DS1, and KIR2DS5.
The antibody was conjugated to BD Horizon™ BUV496 which is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This dye is a tandem fluorochrome of BD Horizon BUV395 with an Ex Max of 348-nm and an acceptor dye with an Em Max at 496-nm. BD Horizon BUV496 can be excited by the ultraviolet laser (355 nm) and detected with a 515/30 nm filter with a 450LP. Due to the excitation of the acceptor dye by other laser lines, there may be significant spillover into the channel detecting BD Horizon V500 or BV510 (eg, 525/40-nm filter). However, the spillover can be corrected through compensation as with any other dye combination.
Development References (9)
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Estefanía E, Flores R, Gómez-Lozano N, Aguilar H, López-Botet M, Vilches C. Human KIR2DL5 is an inhibitory receptor expressed on the surface of NK and T lymphocyte subsets.. J Immunol. 2007; 178(7):4402-10. (Clone-specific: Flow cytometry, Immunoprecipitation). View Reference
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Huard B, Prigent P, Pagès F, Bruniquel D, Triebel F. T cell major histocompatibility complex class II molecules down-regulate CD4+ T cell clone responses following LAG-3 binding. Eur J Immunol. 1996; 26(5):1180-1186. (Biology). View Reference
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Ikeda MA, Jakoi L, Nevins JR. A unique role for the Rb protein in controlling E2F accumulation during cell growth and differentiation. Proc Natl Acad Sci U S A. 1996; 93(8):3215-3220. (Biology). View Reference
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Middleton D, Gonzelez F. The extensive polymorphism of KIR genes. Immunol. 2010; 129(1):8-19. (Biology). View Reference
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Pende D, Falco M, Vitale M, et al. Killer Ig-Like Receptors (KIRs): Their Role in NK Cell Modulation and Developments Leading to Their Clinical Exploitation. Front Immunol. 2019; 10:1179. (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.
