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BV605 Mouse Anti-Human HLA-E
BV605 Mouse Anti-Human HLA-E

Multiparameter flow cytometric analysis of HLA-E expression on human peripheral blood leucocyte populations. Human whole blood was stained with either BD Horizon™ BV605 IgG1, κ Isotype Control (Cat. No. 562652; Left Plot) or BD Horizon™ BV605 Mouse Anti-Human HLA-E antibody (Cat. No. 567431/567430; Right Plot). The erythrocytes were lysed with BD FACS™ Lysing Solution (Cat. No. 349202). A two-parameter pseudocolor density plot showing the correlated expression of HLA-E (or Ig Isotype control staining) versus side-light scatter (SSC-A) signals was derived from gated events with the forward and side-light scatter characteristics of intact leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software.

Multiparameter flow cytometric analysis of HLA-E expression on human peripheral blood leucocyte populations. Human whole blood was stained with either BD Horizon™ BV605 IgG1, κ Isotype Control (Cat. No. 562652; Left Plot) or BD Horizon™ BV605 Mouse Anti-Human HLA-E antibody (Cat. No. 567431/567430; Right Plot). The erythrocytes were lysed with BD FACS™ Lysing Solution (Cat. No. 349202). A two-parameter pseudocolor density plot showing the correlated expression of HLA-E (or Ig Isotype control staining) versus side-light scatter (SSC-A) signals was derived from gated events with the forward and side-light scatter characteristics of intact leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software.

Product Details
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BD Horizon™
HLA class I histocompatibility antigen, alpha chain E; HLAE; MHC Class I Antigen E
Human (QC Testing)
Mouse BALB/c IgG1, κ
Human HLA-E Recombinant Protein
Flow cytometry (Routinely Tested)
5 µl
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO


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 to the dye under optimum conditions and unconjugated antibody and free dye were removed.

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).

Product Notices

  1. 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).
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  4. 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.
  5. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  6. Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
  7. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  8. CF™ is a trademark of Biotium, Inc.
  9. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from BD Horizon™ BV421 may be observed. Therefore, we recommend that individual compensation controls be performed for every BD Horizon™ BV605 conjugate.
  10. BD Horizon Brilliant Violet 605 is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,227,187; 8,455,613; 8,575,303; 8,354,239.
  11. 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.
  12. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
567431 Rev. 1
Antibody Details
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3D12

The 3D12 monoclonal antibody specifically recognizes Human Leukocyte Antigen E (HLA-E) that is widely expressed on leucocytes and some other cell types. Cell surface HLA-E is normally expressed as a noncovalent complex comprised of the ~45 kDa type I transmembrane, HLA-E heavy-chain glycoprotein, the ~12 kDa invariant β2-microglobulin (β2m) light chain, and a short bound peptide. Human HLA-E represents a nonclassical Major Histocompatibility Complex class I (MHC class Ib) molecule that is homologous to mouse H-2 Qa-1. Although structurally related to the classical, highly polymorphic MHC class Ia antigens (HLA-A, -B, -C), HLA-E shows limited polymorphism. HLA-E functions in the regulation or self-nonself discrimination of innate and adaptive immune responses. In addition to binding self peptides, the HLA-E complex can selectively bind and present peptides derived from bacterial or viral pathogen-infected cells, stressed cells, or tumor cells to elicit antigen-specific, HLA-E-restricted CD8+ T cell responses. The cell surface HLA-E complex likewise serves as a ligand for heterodimeric CD94:NKG2A inhibitory and CD94:NKG2C activating receptors that are differentially expressed on NK cells and some T cells. These ligand:receptor interactions can either suppress or promote NK or T cell-mediated responses. The 3D12 antibody reportedly binds to both free or complexed HLA-E heavy chain and can block HLA-E-dependent function.

This antibody is conjugated to BD Horizon™ BV605 which is part of the BD Horizon Brilliant™ Violet family of dyes. With an Ex Max of 407-nm and Em Max of 602-nm, BD Horizon™ BV605 can be excited by a violet laser and detected with a standard 610/20-nm filter set. BD Horizon™ BV605 is a tandem fluorochrome of BD Horizon™ BV421 and an acceptor dye with an Em max at 605-nm. Due to the excitation of the acceptor dye by the green (532 nm) and yellow-green (561 nm) lasers, there will be significant spillover into the PE and BD Horizon™ PE-CF594 detectors off the green or yellow-green lasers. BD Horizon™ BV605 conjugates are very bright, often exhibiting brightness equivalent to PE conjugates and can be used as a third color off of the violet laser.

567431 Rev. 1
Format Details
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BV605
The BD Horizon Brilliant Violet™ 605 (BV605) dye is part of the BD Horizon Brilliant Violet™ family of dyes. This tandem fluorochrome is comprised of a BV421 donor with an excitation maximum (Ex Max) of 407-nm and an acceptor dye with an emission maximum (Em Max) at 605-nm. BV605, driven by BD innovation, is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 610-nm (e.g., a 610/20-nm bandpass filter). The acceptor dye can be excited by the yellow-green (561-nm) laser resulting in cross-laser excitation and fluorescence spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BV605
Violet 405 nm
407 nm
605 nm
567431 Rev.1
Citations & References
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Development References (6)

  1. Joosten SA, Sullivan LC, Ottenhoff TH. Characteristics of HLA-E Restricted T-Cell Responses and Their Role in Infectious Diseases. J Immunol Res. 2016; 2016:1-11. (Biology). View Reference
  2. Joosten SA, van Meijgaarden KE, van Weeren PC, et al. Mycobacterium tuberculosis peptides presented by HLA-E molecules are targets for human CD8 T-cells with cytotoxic as well as regulatory activity.. PLoS Pathog. 2010; 6(2):e1000782. (Clone-specific: Flow cytometry). View Reference
  3. Lee N, Goodlett DR, Ishitani A, Marquardt H, Geraghty DE. HLA-E surface expression depends on binding of TAP-dependent peptides derived from certain HLA class I signal sequences.. J Immunol. 1998; 160(10):4951-60. (Immunogen: Flow cytometry, Immunoprecipitation, Western blot). View Reference
  4. Lee N, Llano M, Carretero M, et al. HLA-E is a major ligand for the natural killer inhibitory receptor CD94/NKG2A.. Proc Natl Acad Sci USA. 1998; 95(9):5199-204. (Clone-specific: Functional assay, Immunoprecipitation, Western blot). View Reference
  5. Llano M, Lee N, Navarro F, et al. HLA-E-bound peptides influence recognition by inhibitory and triggering CD94/NKG2 receptors: preferential response to an HLA-G-derived nonamer.. Eur J Immunol. 1998; 28(9):2854-63. (Clone-specific: Flow cytometry, Functional assay). View Reference
  6. Ogg G, Cerundolo V, McMichael AJ. Capturing the antigen landscape: HLA-E, CD1 and MR1.. Curr Opin Immunol. 2019; 59:121-129. (Biology). View Reference
View All (6) View Less
567431 Rev. 1

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.

Refer to manufacturer's instructions for use and related User Manuals and Technical Data Sheets before using this product 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.