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PerCP-Cy™5.5 Rat Anti-Mouse I-A/I-E

BD Pharmingen™ PerCP-Cy™5.5 Rat Anti-Mouse I-A/I-E

Clone M5/114.15.2 (also known as M5/114)

(RUO)
PerCP-Cy™5.5 Rat Anti-Mouse I-A/I-E
Multicolor flow cytometric analysis of I-A/I-E MHC class II alloantigen expression on splenocytes from positive and negative mouse strains. Mouse spleen cells from either M5/114-negative SJL (Left Panel) or M5/114-positive BALB/c (Right Panel) mice were stained with PerCP-Cy™5.5 Rat Anti-Mouse I-A/I-E (Cat. No. 562363), APC Rat Anti-Mouse CD45R/B220 (Cat. No. 553092/561880) and APC Rat Anti-Mouse CD11b (Cat. No. 553312/561690). Two-color flow cytometric dot plots showing the expression of I-A/I-E MHC class II alloantigens versus CD45R/B220 and CD11b were derived from gated events with the forward and side light-scatter characteristics of viable splenocytes. The M5/114 monoclonal antibody detects I-Ad and I-Ed MHC class II alloantigens that are expressed on both B cells and macrophages. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Multicolor flow cytometric analysis of I-A/I-E MHC class II alloantigen expression on splenocytes from positive and negative mouse strains. Mouse spleen cells from either M5/114-negative SJL (Left Panel) or M5/114-positive BALB/c (Right Panel) mice were stained with PerCP-Cy™5.5 Rat Anti-Mouse I-A/I-E (Cat. No. 562363), APC Rat Anti-Mouse CD45R/B220 (Cat. No. 553092/561880) and APC Rat Anti-Mouse CD11b (Cat. No. 553312/561690). Two-color flow cytometric dot plots showing the expression of I-A/I-E MHC class II alloantigens versus CD45R/B220 and CD11b were derived from gated events with the forward and side light-scatter characteristics of viable splenocytes. The M5/114 monoclonal antibody detects I-Ad and I-Ed MHC class II alloantigens that are expressed on both B cells and macrophages. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
Product Details
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BD Pharmingen™
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_11153297
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 with PerCP-Cy5.5 under optimum conditions, and unconjugated antibody and free PerCP-Cy5.5 were removed. Storage of PerCP-Cy5.5 conjugates in unoptimized diluent is not recommended and may result in loss of signal intensity.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Cy is a trademark of Amersham Biosciences Limited. This conjugated product is sold under license to the following patents: US Patent Nos. 5,486,616; 5,569,587; 5,569,766; 5,627,027.
  4. 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.
  5. 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.
  6. PerCP-Cy5.5–labelled antibodies can be used with FITC- and R-PE–labelled reagents in single-laser flow cytometers with no significant spectral overlap of PerCP-Cy5.5, FITC, and R-PE fluorescence.
  7. PerCP-Cy5.5 is optimized for use with a single argon ion laser emitting 488-nm light. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using dual-laser cytometers, which may directly excite both PerCP and Cy5.5™. We recommend the use of cross-beam compensation during data acquisition or software compensation during data analysis.
  8. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  9. This product is subject to proprietary rights of Amersham Biosciences Corp. and Carnegie Mellon University and made and sold under license from Amersham Biosciences Corp. This product is licensed for sale only for research. It is not licensed for any other use. If you require a commercial license to use this product and do not have one return this material, unopened to BD Biosciences, 10975 Torreyana Rd, San Diego, CA 92121 and any money paid for the material will be refunded.
  10. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
562363 Rev. 2
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.

562363 Rev. 2
Format Details
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PerCP-Cy5.5
PerCP-Cy5.5 dye is part of the BD blue family of dyes. This tandem fluorochrome is comprised of a fluorescent protein complex (PerCP) with an excitation maximum (Ex Max) of 482 nm and an acceptor dye with an emission maximum (Em Max) at 676 nm. PerCP-Cy5 is designed to be excited by the blue laser (488-nm) and detected using an optical filter centered near 680 nm (e.g., a 695/40 nm bandpass filter). The donor dye can be partially excited by the Violet (405-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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PerCP-Cy5.5
Blue 488 nm
482 nm
676 nm
562363 Rev.2
Citations & References
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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. 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
  3. 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: Flow cytometry). View Reference
  4. 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: Immunofluorescence, Immunohistochemistry). View Reference
  5. Shi Y, Kaliyaperumal A, Lu L, et al. Promiscuous presentation and recognition of nucleosomal autoepitopes in lupus: role of autoimmune T cell receptor alpha chain. J Exp Med. 1998; 187(3):367-378. (Clone-specific: Blocking). 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
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562363 Rev. 2

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For Research Use Only. Not for use in diagnostic or therapeutic procedures.