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PE Rat Anti-Mouse CD223
PE Rat Anti-Mouse CD223
The expression of CD223 on activated T lymphocytes. C57BL/6 splenocytes were activated by culture for 2 days in the presence of immobilized anti-mouse CD3e mAb 145-2C11 (Cat. No. 553057) and then were stained with FITC-conjugated anti-mouse CD3e mAb 145-2C11 (Cat. No. 553061/553062) and either PE-conjugated rat IgG1, κ isotype control mAb R3-34 (Cat. No. 553925, left panel) or PE-conjugated mAb C9B7W (right panel) in the presence of Mouse BD Fc Block™ purified anti-mouse CD16/CD32, mAb 2.4G2 (Cat. No. 553141/553142). Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.
The expression of CD223 on activated T lymphocytes. C57BL/6 splenocytes were activated by culture for 2 days in the presence of immobilized anti-mouse CD3e mAb 145-2C11 (Cat. No. 553057) and then were stained with FITC-conjugated anti-mouse CD3e mAb 145-2C11 (Cat. No. 553061/553062) and either PE-conjugated rat IgG1, κ isotype control mAb R3-34 (Cat. No. 553925, left panel) or PE-conjugated mAb C9B7W (right panel) in the presence of Mouse BD Fc Block™ purified anti-mouse CD16/CD32, mAb 2.4G2 (Cat. No. 553141/553142). Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.
Product Details
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BD Pharmingen™
Lag3; LAG-3; Lymphocyte-activation gene 3; Ly66
Mouse (QC Testing)
Rat LEW, also known as Lewis IgG1, κ
Mouse LAG3 fusion protein
Flow cytometry (Routinely Tested)
0.2 mg/ml
AB_394374
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


Preparation And Storage

The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with R-PE under optimum conditions, and unconjugated antibody and free PE were removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.

Recommended Assay Procedures

This antibody conjugate has been tested by immunofluorescent staining and flow cytometry. We recommend the use of Mouse BD Fc Block™ purified anti-mouse CD16/CD32 mAb 2.4G2 (Cat. No. 553141/553142) to reduce nonspecific binding of the PE conjugate to cells bearing Fcγ receptors. Other reported applications for this clone, include in vitro and in vivo inhibition of CD223 function, for use in these applications, use our no azide/low endotoxin format (NA/LE™), Cat. No. 552379.

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Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  3. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  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.
552380 Rev. 4
Antibody Details
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C9B7W

The C9B7W antibody monoclonal antibody specifically binds to an epitope in the D2 domain of CD223 (LAG3), the 70-kDa protein encoded by Lymphocyte-activation gene 3 (Lag3). A fusion protein consisting of the entire extracellular region of mouse LAG3 with mouse IgG1 was used as immunogen. CD223 is a type-I membrane protein with four extracellular Ig-like domains; it is structurally homologous to CD4; and, like CD4, it binds MHC class II molecules. However, unlike CD4, it is not expressed on resting human and mouse T lymphocytes. In the mouse, as previously described in the human, CD223 expression is upregulated on T lymphocytes (both CD4+ and CD8+) activated through the T-cell receptor (TCR) and on IL-2-activated NK (LAK) cells, and it is not detected on B cells, dendritic cells, or Phorbol 12-myristate 13-acetate (PMA)-stimulated splenocytes. Studies on human peripheral T lymphocytes suggest that CD223 associates with the TCR to downregulate TCR signaling. In contrast, in vivo and in vitro evaluations of vaccination protocols in mice suggest that CD223 promotes immune responses by activating antigen-presenting cells. Furthermore, NK cells of Lag3-/- mice display defects in their capacity to kill certain tumor cells. Mouse CD223 also has been demonstrated to contribute to the suppressor function of T regulatory cells and the C9B7W antibody has been shown to inhibit this function in vitro and in vivo. Therefore, CD223 appears to play complex roles in the regulation of immune responses. Although the C9B7W antibody is unable to block the binding of MHC class II-IgG2a fusion protein to CD223, it is able to block the CD223-mediated inhibition of IL-2 production by a T-cell hybridoma responding to antigen.

552380 Rev. 4
Format Details
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PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
552380 Rev.4
Citations & References
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Development References (10)

  1. Baixeras E, Huard B, Miossec C, et al. Characterization of the lymphocyte activation gene 3-encoded protein. A new ligand for human leukocyte antigen class II antigens. J Exp Med. 1992; 176(2):327-337. (Biology). View Reference
  2. El Mir S, Triebel F. A soluble lymphocyte activation gene-3 molecule used as a vaccine adjuvant elicits greater humoral and cellular immune responses to both particulate and soluble antigens. J Immunol. 2000; 164(11):5583-5589. (Biology). View Reference
  3. Hannier S, Tournier M, Bismuth G, Triebel F. CD3/TCR complex-associated lymphocyte activation gene-3 molecules inhibit CD3/TCR signaling. J Immunol. 1998; 161(8):4058-4065. (Biology). View Reference
  4. Hannier S, Triebel F. The MHC class II ligand lymphocyte activation gene-3 is co-distributed with CD8 and CD3-TCR molecules after their engagement by mAb or peptide-MHC class I complexes. Int Immunol. 1999; 11(11):1745-1752. (Biology). View Reference
  5. Huang CT, Workman CJ, Flies D, et al. Role of LAG-3 in regulatory T cells. Immunity. 2004; 21(4):503-513. (Clone-specific: Inhibition). View Reference
  6. Huard B, Mastrangeli R, Prigent P, et al. Characterization of the major histocompatibility complex class II binding site on LAG-3 protein. Proc Natl Acad Sci U S A. 1997; 94(11):5744-5749. (Biology). View Reference
  7. Miyazaki T, Dierich A, Benoist C, Mathis D. Independent modes of natural killing distinguished in mice lacking Lag3. Science. 1996; 272(5260):405-408. (Biology). View Reference
  8. Prigent P, El Mir S, Dréano M, Triebel F. Lymphocyte activation gene-3 induces tumor regression and antitumor immune responses. Eur J Immunol. 1999; 29(12):3867-3876. (Biology). View Reference
  9. Triebel F, Jitsukawa S, Baixeras E, et al. LAG-3, a novel lymphocyte activation gene closely related to CD4. J Exp Med. 1990; 171(5):1393-1405. (Biology). View Reference
  10. Workman CJ, Rice DS, Dugger KJ, Kurschner C, Vignali DA. Phenotypic analysis of the murine CD4-related glycoprotein, CD223 (LAG-3). Eur J Immunol. 2002; 32(8):2255-2263. (Immunogen: Inhibition). View Reference
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552380 Rev. 4

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.