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This product is the replacement for 748845.
569176Image1.png

Multicolor flow cytometric analysis of I-A/I-E MHC Class II alloantigen expression on viable Mouse splenic leucocytes. C57BL/6 Mouse splenocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) [Cat. No. 553141/553142]. The cells were then stained with PerCP-Cy5.5 Hamster Anti-Mouse CD3e antibody (Cat. No. 551163/561108) and with either BD Horizon™ BUV737 Rat IgG2b, κ Isotype Control (Cat. No. 612762; Left Plot) or BD Horizon™ BUV737 Rat Anti-Mouse I-A/I-E antibody (Cat. No. 569176; Right Plot) at 0.25 µg/test. DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride) Solution (Cat. No. 564907) was added to cells right before analysis. The bivariate pseudocolor density plot showing the correlated expression of I-A/I-E (or Ig Isotype control staining) versus CD3e was derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

569176-2.png

Multicolor flow cytometric analysis of I-A/I-E MHC Class II alloantigen expression on viable Mouse splenic leucocytes. C57BL/6 Mouse splenocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) [Cat. No. 553141/553142]. The cells were then stained with PerCP-Cy5.5 Hamster Anti-Mouse CD3e antibody (Cat. No. 551163/561108) and with either BD Horizon™ BUV737 Rat IgG2b, κ Isotype Control (Cat. No. 612762; Left Plot) or BD Horizon™ BUV737 Rat Anti-Mouse I-A/I-E antibody (Cat. No. 569176; Right Plot) at 0.25 µg/test. DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride) Solution (Cat. No. 564907) was added to cells right before analysis. The bivariate pseudocolor density plot showing the correlated expression of I-A/I-E (or Ig Isotype control staining) versus CD3e was derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

569176Image1.png 569176-2.png Orange-Cap_UV.png

Multicolor flow cytometric analysis of I-A/I-E MHC Class II alloantigen expression on viable Mouse splenic leucocytes. C57BL/6 Mouse splenocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) [Cat. No. 553141/553142]. The cells were then stained with PerCP-Cy5.5 Hamster Anti-Mouse CD3e antibody (Cat. No. 551163/561108) and with either BD Horizon™ BUV737 Rat IgG2b, κ Isotype Control (Cat. No. 612762; Left Plot) or BD Horizon™ BUV737 Rat Anti-Mouse I-A/I-E antibody (Cat. No. 569176; Right Plot) at 0.25 µg/test. DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride) Solution (Cat. No. 564907) was added to cells right before analysis. The bivariate pseudocolor density plot showing the correlated expression of I-A/I-E (or Ig Isotype control staining) versus CD3e was derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

Multicolor flow cytometric analysis of I-A/I-E MHC Class II alloantigen expression on viable Mouse splenic leucocytes. C57BL/6 Mouse splenocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) [Cat. No. 553141/553142]. The cells were then stained with PerCP-Cy5.5 Hamster Anti-Mouse CD3e antibody (Cat. No. 551163/561108) and with either BD Horizon™ BUV737 Rat IgG2b, κ Isotype Control (Cat. No. 612762; Left Plot) or BD Horizon™ BUV737 Rat Anti-Mouse I-A/I-E antibody (Cat. No. 569176; Right Plot) at 0.25 µg/test. DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride) Solution (Cat. No. 564907) was added to cells right before analysis. The bivariate pseudocolor density plot showing the correlated expression of I-A/I-E (or Ig Isotype control staining) versus CD3e was derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) leucocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

Product Details
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BD Horizon™
H-2I; I-Ab, I-Ad, I-Aq, I-Ed, and I-Ek MHC class II alloAgs; Ia Ag; M5/114; MHC II
Mouse (QC Testing)
Rat BN x LEW IgG2b, κ
Activated C57BL/6 Mouse Spleen Cells
Flow cytometry (Routinely Tested)
0.2 mg/ml
111861
AB_3684842
Aqueous buffered solution containing ≤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.
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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® CompBeads 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).

   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).

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Product Notices

  1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  2. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  3. An isotype control should be used at the same concentration as the antibody of interest.
  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. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  6. BD Horizon Brilliant Ultraviolet 737 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.
  7. 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.
  8. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  9. For U.S. patents that may apply, see bd.com/patents.
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569176 Rev. 1
Antibody Details
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M5/114.15.2

The M5/114.15.2 monoclonal antibody recognizes a polymorphic determinant shared by the I-A[b], I-A[d], I-A[q], I-E[d], and I-E[k] (but not I-A[f], I-A[k], or I-A[s]) MHC class II alloantigens that can be expressed by B cells, dendritic cells, monocytes, macrophages and activated T cells. It also reacts with cells from mice of the H-2[p] and H-2[r] haplotypes, and it is non-reactive with cells from NOD (H-2[g7]) mice. Flow cytometric analysis indicates that the M5/114.15.2 and 2G9 monoclonal antibodies have comparable reactivity on cells from mice with I-A[b], I-A[d], I-A[g7], I-A[q], I-E[d], and I-E[k] alloantigens.

569176 Rev. 1
Format Details
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BUV737
The BD Horizon Brilliant™ Ultraviolet 737 (BUV737) Dye is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This tandem fluorochrome is comprised of a BUV395 donor with an excitation maximum (Ex Max) of 350-nm and an acceptor dye with an emission maximum (Em Max) at 735-nm. BUV737, driven by BD innovation, is designed to be excited by the ultraviolet laser (355-nm) and detected using an optical filter centered near 740-nm (e.g., 740/35 bandpass filter). The acceptor dye can be excited by the Red (628–640nm) 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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BUV737
350 nm
735 nm
569176 Rev.1
Citations & References
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View product citations for antibody "569176" on CiteAb

Development References (7)

  1. Bhattacharya A, Dorf ME, Springer TA. A shared alloantigenic determinant on Ia antigens encoded by the I-A and I-E subregions: evidence for I region gene duplication. J Immunol. 1981; 127(6):2488-2495. (Immunogen: Immunoprecipitation). View Reference
  2. Ernst DN, McQuitty DN, Weigle WO, Hobbs MV. Expression of membrane activation antigens on murine B lymphocytes stimulated with lipopolysaccharide. Cell Immunol. 1988; 114(1):161-173. (Clone-specific: Flow cytometry). View Reference
  3. Guo MW, Watanabe T, Mori E, Mori T. Molecular structure and function of CD4 on murine egg plasma membrane. Zygote. 1995; 3(1):65-73. (Clone-specific: Blocking). View Reference
  4. Hattori M, Buse JB, Jackson RA, et al. The NOD mouse: recessive diabetogenic gene in the major histocompatibility complex. Science. 1986; 231(4739):733-735. (Clone-specific). View Reference
  5. Nelson AJ, Hosier S, Brady W, Linsley PS, Farr AG. Medullary thymic epithelium expresses a ligand for CTLA4 in situ and in vitro. J Immunol. 1998; 151(5):2453-2461. (Clone-specific: Blocking, Immunofluorescence, Immunohistochemistry). View Reference
  6. Viville S, Neefjes J, Lotteau V, et al. Mice lacking the MHC class II-associated invariant chain. Cell. 1993; 72(4):635-648. (Clone-specific: Flow cytometry, Immunofluorescence). View Reference
  7. Yamashita I, Nagata T, Tada T, Nakayama T. CD69 cell surface expression identifies developing thymocytes which audition for T cell antigen receptor-mediated positive selection. Int Immunol. 1993; 5(9):1139-1150. (Clone-specific: Blocking). View Reference
View All (7) View Less
569176 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.