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Multicolor flow cytometric analysis of CD366 (TIM-3) expression on viable activated Mouse splenic leukocytes. BALB/c Mouse splenocytes were cultured for 4 days in the presence of plate-bound Purified NA/LE Hamster Anti-Mouse CD3e (Cat. No. 553057/567114; 10 μg/ml for coating), soluble Purified NA/LE Hamster Anti-Mouse CD28 (Cat. No. 567110/553294; 2 μg/ml), and Purified NA/LE Rat Anti-Mouse IL-4 (Cat. No. 554432) antibodies. The cells were harvested, washed and preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) [Cat. No. 553141/553142]. The cells were then stained with PE Rat Anti-Mouse CD8a antibody (Cat. No. 553032) and with either BD Horizon™ R718 Rat IgG2a, κ Isotype Control (Cat. No. 566941; Left Plot) or BD Horizon™ R718 Rat Anti-Mouse CD366 (TIM-3) antibody (Cat. No. 569809/569883; Right Plot) at 0.5 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 expression of CD366 (TIM-3) [or Ig Isotype control staining] versus CD8a was derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) splenic lymphocytes. 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.
BD Horizon™ R718 Rat Anti-Mouse CD366 (TIM-3)
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The 25F.1D6 monoclonal antibody specifically recognizes CD366 which is also known as TIM-3 (T-cell immunoglobulin and mucin domain-containing 3, T-cell immunoglobulin mucin receptor 3, or T-cell membrane protein 3). CD366 (TIM-3) is an ~31 kDa type I transmembrane glycoprotein encoded by Havcr2 (Hepatitis A virus cellular receptor 2) that belongs to the TIM family within the Ig superfamily. CD366 (TIM-3) is comprised of one IgV-type domain followed by a serine/threonine-rich mucin stalk region in its extracellular region, a transmembrane segment, and a tyrosine phosphorylation motif in its cytoplasmic tail. CD366 (TIM-3) expression is upregulated on subpopulations of activated myeloid cells including macrophages, monocytes, dendritic cells (DC), microglia, mast cells as well as on Type-1 CD4+ (Th1-like) T cells, cytotoxic CD8+ T cells, regulatory T cells (Treg), and natural killer (NK) cells. CD366 (TIM-3) functions as an inhibitory receptor that helps maintain immunological homeostasis and self-tolerance. It may also serve an immune checkpoint molecule that inhibits antitumor immunity and promotes T cell exhaustion. Crosslinking of cell surface CD366 (TIM-3) by Galectin-9 binding downregulates Th1-like and CD8+ T cell responses and can promote Treg or myeloid-derived suppressor cells. CD366 (TIM-3) enables DC to bind phosphatidyl serine expressed by apoptotic cells, phagocytize these cells to suppress inflammation and promote antigen cross-presentation. CD366 (TIM-3) can also bind to high mobility group protein B1 (HMGB1) and inhibit stimulation of the immune response to nucleic acids released by dying tumor cells.
Development References (9)
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Anderson AC, Joller N, Kuchroo VK. Lag-3, Tim-3, and TIGIT: Co-inhibitory Receptors with Specialized Functions in Immune Regulation.. Immunity. 2016; 44(5):989-1004. (Biology). View Reference
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Chiba S, Baghdadi M, Akiba H, et al. Tumor-infiltrating DCs suppress nucleic acid-mediated innate immune responses through interactions between the receptor TIM-3 and the alarmin HMGB1.. Nat Immunol. 2012; 13(9):832-42. (Biology). View Reference
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Monney L1, Sabatos CA, Gaglia JL, et al. Th1-specific cell surface protein Tim-3 regulates macrophage activation and severity of an autoimmune disease.. Nature. 2002; 415(6871):536-541. (Clone-specific: Flow cytometry). View Reference
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Nakae S, Iikura M, Suto H, et al. TIM-1 and TIM-3 enhancement of Th2 cytokine production by mast cells.. Blood. 2007; 110(7):2565-8. (Biology). View Reference
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Nakayama M, Akiba H, Takeda K, et al. Tim-3 mediates phagocytosis of apoptotic cells and cross-presentation. Blood. 2009; 113(16):3821-3830. (Biology). View Reference
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Oikawa T1, Kamimura Y, Akiba H, et al. Preferential involvement of Tim-3 in the regulation of hepatic CD8+ T cells in murine acute graft-versus-host disease.. J Immunol. 2006; 177(7):4281-4287. (Biology). View Reference
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Phong BL, Avery L, Sumpter TL, et al. Tim-3 enhances FcεRI-proximal signaling to modulate mast cell activation.. J Exp Med. 2015; 212(13):2289-304. (Biology). View Reference
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Veenstra RG, Taylor PA, Zhou Q, et al. Contrasting acute graft-versus-host disease effects of Tim-3/galectin-9 pathway blockade dependent upon the presence of donor regulatory T cells.. Blood. 2012; 120(3):682-90. (Biology). View Reference
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Zhu C, Anderson AC, Schubart A, et al. The Tim-3 ligand galectin-9 negatively regulates T helper type 1 immunity.. Nat Immunol. 2005; 6(12):1245-52. (Biology). View Reference
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.