TIM-3 Antibody [Unconjugated]

TIM-3 mRNA was detected in formalin-fixed paraffin-embedded tissue sections of human tonsil probed with ACD RNAScope® Probe (Catalog # 560681) and stained using ACD RNAscope® 2.5 HD Detection Reagents-Red (top image, Catalog # 32260). Adjacent tissue section was processed for immunohistochemistry using R&D Systems Goat Anti-Human TIM-3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2365) at 3 ug/mL for 1 hour at room temperature followed by incubation with the Anti-Mouse IgG VisUCyte HRP Polymer Antibody (R&D Systems, Catalog # VC004) and DAB chromogen (lower image, yellow-brown). Tissues were counterstained with hematoxylin (blue).

TIM‑3 was detected in immersion fixed paraffin-embedded sections of human tonsil using Goat Anti-Human TIM‑3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2365) at 3 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC004). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to cell membranes and extracellular space. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Western blot shows lysates of HDLM‑2 human Hodgkin’s lymphoma cell line and human TIM-3 knockout HDLM‑2 human Hodgkin’s lymphoma cell line (KO). PVDF membrane was probed with 1 µg/mL of Goat Anti-Human TIM‑3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2365) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF017). A specific band was detected for TIM‑3 at approximately 50 kDa (as indicated) in the parental HDLM‑2 human Hodgkin’s lymphoma cell line, but is not detectable in knockout HDLM‑2 human Hodgkin’s lymphoma cell line. GAPDH (AF5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Western Blot Buffer Group 1.

Immunohistochemical staining of peritoneum tissue for CD4, CD8, CD56, OPN, CD68, Gal-9, and TIM-3. HE, hematoxylin and eosin. Magnification, 20×. Arrows indicate loosen tissue at the broken boundary of the granuloma. Image collected and cropped by CiteAb from the following publication (//www.mdpi.com/1422-0067/18/7/1382), licensed under a CC-BY license. Not internally tested by R&D Systems.

TIM-3 Association with Intracellular Kinases in CD8+/MART-1+ T cells.TIM-3 co-immunoprecipitation analysis of unactivated and 15 min. stimulation with anti-CD3/CD28 beads (activated). Equivalent amounts of protein (~2mg) were co-immunoprecipitated with pAb anti-TIM-3 antibody and western blot was performed using capillary electrophoresis. Cleared lysate served as a loading control for individual antibody reactivity. Image collected and cropped by CiteAb from the following publication (//dx.plos.org/10.1371/journal.pone.0140694), licensed under a CC-BY license. Not internally tested by R&D Systems.

• Reactivity: Hu
• Applications: WB, Flow, IHC, CyTOF-ready, Dual ISH-IHC, KO
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Unconjugated
• Concentration: LYOPH

TIM-3 Antibody [Unconjugated] Summary

• Immunogen: Mouse myeloma cell line NS0-derived recombinant human TIM‑3
  Ser22-Arg200
  Accession # Q8TDQ0
• Specificity: Detects human TIM-3 in direct ELISAs and Western blots. In direct ELISAs, approximately 5% cross-reactivity with recombinant mouse (rm) TIM-3 is observed and less than 1% cross-reactivity with recombinant human TIM-1, rmTIM-1, and rmTIM-2 is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Gene: HAVCR2
• Purity Statement: Antigen Affinity-purified

Applications/Dilutions

• Dilutions:
  • CyTOF-ready
  • Dual RNAscope ISH-IHC 3-25 ug/mL
  • Flow Cytometry 0.25 ug/10^6 cells
  • Immunohistochemistry 3-15 ug/mL
  • Knockout Validated 1 ug/mL
  • Western Blot 0.1 ug/mL

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  12 months from date of receipt, -20 to -70 degreesC as supplied. 1 month, 2 to 8 degreesC under sterile conditions after reconstitution. 6 months, -20 to -70 degreesC under sterile conditions after reconstitution.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.2 mg/mL in sterile PBS.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for TIM-3 Antibody [Unconjugated]

• CD366
• FLJ14428
• HAVCR2
• HAVcr-2
• hepatitis A virus cellular receptor 2
• kidney injury molecule-3
• KIM-3
• SPTCL
• T cell immunoglobulin mucin 3
• T cell immunoglobulin mucin-3
• T-cell immunoglobulin and mucin domain-containing protein 3
• TIM 3
• TIM3 T-cell membrane protein 3
• TIM3
• TIM-3
• TIMD3
• TIMD-3
• TIMD3KIM-3

Background

TIM-3 (T cell immunoglobulin and mucin domain-3) is a 60 kDa member of the TIM family of immune regulating molecules. TIMs are type I transmembrane glycoproteins with one Ig-like V-type domain and a Ser/Thr-rich mucin stalk (1-3). There are three TIM genes in human and eight in mouse. Mature human TIM-3 consists of a 181 amino acid (aa) extracellular domain (ECD), a 21 aa transmembrane segment, and a 78 aa cytoplasmic tail (4). An alternately spliced isoform is truncated following a short substitution after the Ig-like domain. Within the ECD, human TIM-3 shares 58% aa sequence identity with mouse and rat TIM-3. TIM-3 is expressed on the surface of effector T cells (CD4 + Th1 and CD8 + Tc1) but not on helper T cells (CD4 + Th2 and CD8 + Tc2) (4, 5). In chronic inflammation, autoimmune disorders, and some cancers, TIM-3 is upregulated on several other hematopoietic cell types. It also occurs on hippocampal neurons (7-10). The Ig domain of TIM-3 interacts with a ligand on resting but not activated Th1 and Th2 cells (5, 11). The glycosylated Ig domain of TIM-3 binds cell-associated galectin-9. This induces TIM-3 Tyr phosphorylation and pro-apoptotic signaling (8, 12). TIM-3 functions as a negative regulator of Th1 cell activity. Its blockade results in increased IFN-gamma production, Th1 cell proliferation and cytotoxicity (5, 10, 11, 13), regulatory T cell development (5), and increases in macrophage and neutrophil infiltration into sites of inflammation (14). Soluble mouse TIM-3 constructs which lack the cytoplasmic domain have been shown to inhibit anti-tumor effector T cell responses and to enhance autoimmune reactions (5, 15).

1. Anderson, A.C. and D.E. Anderson (2006) Curr. Opin. Immunol. 18:665.
2. Mariat, C. et al. (2005) Phil. Trans. R. Soc. B. 360:1681.
3. Meyers, J.H. et al. (2005) Trends Mol. Med. 11:362.
4. Monney, L. et al. (2002) Nature 415:536.
5. Sanchez-Fueyo, A. et al. (2003) Nat. Immunol. 4:1093.
6. Khademi, M. et al. (2004) J. Immunol. 172:7169.
7. Wiener, Z. et al. (2007) J. Invest. Dermatol. 127:906.
8. van de Weyer, P.S. et al. (2006) Biochem. Biophys. Res. Commun. 351:571.
9. Gielen, A.W. et al. (2005) J. Neuroimmunol. 164:93.
10. Oikawa, T. et al. (2006) J. Immunol. 177:4281.
11. Sabatos, C.A. et al. (2003) Nat. Immunol. 4:1102.
12. Zhu, C. et al. (2005) Nat. Immunol. 6:1245.
13. Koguchi, K. et al. (2006) J. Exp. Med. 203:1413.
14. Frisancho-Kiss, S. et al. (2006) J. Immunol. 176:6411.
15. Geng, H. et al. (2006) J. Immunol. 176:1411.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for TIM-3 Antibody (AF2365)(36)

Publications using AF2365

• Sánchez-Cerrillo, I;Agudo-Lera, M;Popova, O;Tsukalov, I;Calvet-Mirabent, M;de Los Santos, I;García-Fraile, L;Fuentes, P;Delgado-Arévalo, C;Alcain, J;Sánchez-Gaona, N;Grau-Expósito, J;Lázaro-Díez, M;Muñoz-Calleja, C;Alfranca, A;Genescà, M;Prado, JG;Vrbanac, V;Balazs, A;Buzón, MJ;Toribio, ML;Muñoz-Fernández, MA;Sánchez-Madrid, F;Martín-Gayo, E; Combined dendritic cell and anti-TIGIT immunotherapy potentiates adaptive NK cells against HIV-1 EMBO molecular medicine 2025-06-05 [PMID: 40473838] (Neutralization, Human)
• Jeon, YK;Koh, J;Lee, D;Kim, S;Song, SG;Han, B;Jeong, H;Kim, YA;Keam, B;Lee, SH;Kim, K;Chung, DH; Spatially resolved whole-transcriptomic and proteomic profiling of lung cancer and its immune-microenvironment according to PD-L1 expression Cancer immunology research 2024-09-05 [PMID: 39235761] (Immunohistochemistry, Human)
• Saliby, RM;El Zarif, T;Bakouny, Z;Shah, V;Xie, W;Flippot, R;Denize, T;Kane, MH;Madsen, KN;Ficial, M;Hirsch, L;Wei, XX;Steinarter, JA;Harshman, LC;Vaishampayan, UN;Severgnini, M;McDermott, DF;Lee, GM;Xu, W;Van Allen, EM;McGregor, BA;Signoretti, S;Choueiri, TK;McKay, RR;Braun, DA; Circulating and intratumoral immune determinants of response to atezolizumab plus bevacizumab in patients with variant histology or sarcomatoid renal cell carcinoma Cancer immunology research 2023-06-06 [PMID: 37279009] (IHC, Human)
• M Calvet-Mir, I Sánchez-Ce, N Martín-Cóf, P Martínez-F, H de la Fuen, I Tsukalov, C Delgado-Ar, MJ Calzada, I de Los San, J Sanz, L García-Fra, F Sánchez-Ma, A Alfranca, MÁ Muñoz-Fern, MJ Buzón, E Martín-Gay Antiretroviral therapy duration and immunometabolic state determine efficacy of ex vivo dendritic cell-based treatment restoring functional HIV-specific CD8+ T cells in people living with HIV EBioMedicine, 2022-06-02;81(0):104090. 2022-06-02 [PMID: 35665682] (Neutralization, Human)
• Ziad Bakouny, Ananthan Sadagopan, Praful Ravi, Nebiyou Y. Metaferia, Jiao Li, Shatha AbuHammad et al. Integrative clinical and molecular characterization of translocation renal cell carcinoma Cell Reports 2022-01-04 [PMID: 34986355] (Immunocytochemistry, Human)
• Ziad Bakouny, Ananthan Sadagopan, Praful Ravi, Nebiyou Y. Metaferia, Jiao Li, Shatha AbuHammad et al. Integrative clinical and molecular characterization of translocation renal cell carcinoma Cell Reports 1/4/2022 [PMID: 34986355]
• Courtney M. Smith, Alice Li, Nithya Krishnamurthy, Mark A. Lemmon Phosphatidylserine binding directly regulates TIM-3 function Biochemical Journal 9/17/2021 [PMID: 34435619]
• Courtney M. Smith, Alice Li, Nithya Krishnamurthy, Mark A. Lemmon Phosphatidylserine binding directly regulates TIM-3 function Biochemical Journal 2021-09-17 [PMID: 34435619] (Flow Cytometry, Western Blot, Human)
• Sarradin V, Lusque A, Filleron T et al. Immune microenvironment changes induced by neoadjuvant chemotherapy in triple-negative breast cancers: the MIMOSA-1 study Breast cancer research : BCR 5/26/2021 [PMID: 34039396]
• W Zhong, X Liu, Z Zhu, Q Li, K Li High levels of Tim-3+Foxp3+Treg cells in the tumor microenvironment is a prognostic indicator of poor survival of diffuse large B cell lymphoma patients International immunopharmacology, 2021-04-14;96(0):107662. 2021-04-14 [PMID: 33864956] (IHC, Human)
• David A. Braun, Kelly Street, Kelly P. Burke, David L. Cookmeyer, Thomas Denize, Christina B. Pedersen et al. Progressive immune dysfunction with advancing disease stage in renal cell carcinoma Cancer Cell 5/1/2021 [PMID: 33711273]
• N Ebata, M Fujita, S Sasagawa, K Maejima, Y Okawa, Y Hatanaka, T Mitsuhashi, A Oosawa-Tat, H Tanaka, S Miyano, T Nakamura, S Hirano, H Nakagawa Molecular Classification and Tumor Microenvironment Characterization of Gallbladder Cancer by Comprehensive Genomic and Transcriptomic Analysis Cancers, 2021-02-10;13(4):. 2021-02-10 [PMID: 33578820] (IHC, Human)
• Marina Martinez, Soyeon Kim, Naomi St. Jean, Shaun O’Brien, Lurong Lian, Jing Sun et al. Addition of anti-TIM3 or anti-TIGIT Antibodies to anti-PD1 Blockade Augments Human T cell Adoptive Cell Transfer OncoImmunology 2021-02-04 [PMID: 33537176] (Neutralization, Human)
• Marina Martinez, Soyeon Kim, Naomi St. Jean, Shaun O’Brien, Lurong Lian, Jing Sun et al. Addition of anti-TIM3 or anti-TIGIT Antibodies to anti-PD1 Blockade Augments Human T cell Adoptive Cell Transfer OncoImmunology 2/4/2021 [PMID: 33537176]
• Stéphane Chevrier, Yves Zurbuchen, Carlo Cervia, Sarah Adamo, Miro E. Raeber, Natalie de Souza et al. A distinct innate immune signature marks progression from mild to severe COVID-19 Cell Reports Medicine 1/1/2021 [PMID: 33521697]
• Hui Li, Jinju Wang, Hailing Liu, Tian Lan, Lin Xu, Genshu Wang et al. Existence of intratumoral tertiary lymphoid structures is associated with immune cells infiltration and predicts better prognosis in early-stage hepatocellular carcinoma Aging (Albany NY) 2/22/2020 [PMID: 32087064]
• Wei Li, Jun Han, Kefei Yuan, Hong Wu Integrated tumor stromal features of hepatocellular carcinoma reveals two distinct subtypes with prognostic/predictive significance Aging (Albany NY) 2019-07-12 [PMID: 31299011] (Immunohistochemistry, Human)
• Wei Li, Jun Han, Kefei Yuan, Hong Wu Integrated tumor stromal features of hepatocellular carcinoma reveals two distinct subtypes with prognostic/predictive significance Aging (Albany NY) 7/12/2019 [PMID: 31299011]
• Edna Gordian, Eric A. Welsh, Nicholas Gimbrone, Erin M. Siegel, David Shibata, Ben C. Creelan et al. Transforming growth factor beta -induced epithelial-to-mesenchymal signature predicts metastasis-free survival in non-small cell lung cancer Oncotarget 1/25/2019 [PMID: 30783512]
• DM Franchini, O Lanvin, M Tosolini, E Patras de, A Cammas, S Péricart, CM Scarlata, M Lebras, C Rossi, L Ligat, F Pont, PB Arimondo, C Laurent, M Ayyoub, F Despas, M Lapeyre-Me, S Millevoi, JJ Fournié Microtubule-Driven Stress Granule Dynamics Regulate Inhibitory Immune Checkpoint Expression in T Cells Cell Rep, 2019-01-02;26(1):94-107.e7. 2019-01-02 [PMID: 30605689] (Flow Cytometry, Human)
• D Zhao, M Guo, B Liu, Q Lin, T Xie, Q Zhang, X Jia, Q Shu, X Liang, L Gao, C Ma Frontline Science: Tim-3-mediated dysfunctional engulfment of apoptotic cells in SLE J. Leukoc. Biol., 2017-07-28;0(0):. 2017-07-28 [PMID: 28754800] (Western Blot, Human)
• J Zhao, B Shiratori, H Chagan-Yas, M Matsumoto, T Niki, M Tanaka, Y Takahashi, O Usami, Y Ashino, T Hattori Secretion of IFN-? Associated with Galectin-9 Production by Pleural Fluid Cells from a Patient with Extrapulmonary Tuberculosis Int J Mol Sci, 2017-06-28;18(7):. 2017-06-28 [PMID: 28657598] (IHC, Human)
• TO Mohammed, H Chagan-Yas, Y Ashino, W Nakayama, Y Takahashi, T Shimomura, T Fujimoto, Y Watanabe, T Niki, H Suzushima, T Hattori Galectin-9 as a Predictive Marker for the Onset of Immune-Related Adverse Effects Associated with Anti-CCR4 MoAb Therapy in Patients with Adult T Cell Leukemia Tohoku J. Exp. Med, 2017-03-01;241(3):201-208. 2017-03-01 [PMID: 28321034] (IHC, Human)
• JA Schwartz, KL Clayton, S Mujib, H Zhang, AK Rahman, J Liu, FY Yue, E Benko, C Kovacs, MA Ostrowski Tim-3 is a Marker of Plasmacytoid Dendritic Cell Dysfunction during HIV Infection and Is Associated with the Recruitment of IRF7 and p85 into Lysosomes and with the Submembrane Displacement of TLR9 J. Immunol, 2017-03-06;0(0):. 2017-03-06 [PMID: 28264968] (ICC, Human)
• Guangxiang Luo TIM-1 promotes hepatitis C virus cell attachment and infection J. Virol., 2017-01-03;0(0):. 2017-01-03 [PMID: 27807228] (Western Blot, Human)
• Comprehensive analyses of tumor immunity: implications for cancer immunotherapy Genome Biol, 2016-08-22;17(1):174. 2016-08-22 [PMID: 27549193] (IHC-P, Human)
• Clayton K, Douglas-Vail M, Nur-ur Rahman A, Medcalf K, Xie I, Chew G, Tandon R, Lanteri M, Norris P, Deeks S, Ndhlovu L, Ostrowski M Soluble T cell immunoglobulin mucin domain 3 is shed from CD8+ T cells by the sheddase ADAM10, is increased in plasma during untreated HIV infection, and correlates with HIV disease progression. J Virol, 2015-01-21;89(7):3723-36. 2015-01-21 [PMID: 25609823] (Human)
• Moller-Tank S, Albritton L, Rennert P, Maury W Characterizing functional domains for TIM-mediated enveloped virus entry. J Virol, 2014-04-02;88(12):6702-13. 2014-04-02 [PMID: 24696470] (Flow Cytometry, Human)
• Jiang , Jing, Jin , Mei-Shan, Kong , Fei, Cao , Donghui, Ma , Hong-Xi, Jia , Zhifang, Wang , Yin-Ping, Suo , Jian, Cao , Xueyuan Decreased galectin-9 and increased Tim-3 expression are related to poor prognosis in gastric cancer. PLoS ONE, 2013-12-10;8(12):e81799. 2013-12-10 [PMID: 24339967] (IHC-P, Human)
• Clayton K, Haaland M, Douglas-Vail M, Mujib S, Chew G, Ndhlovu L, Ostrowski M T cell Ig and mucin domain-containing protein 3 is recruited to the immune synapse, disrupts stable synapse formation, and associates with receptor phosphatases. J Immunol, 2013-12-13;192(2):782-91. 2013-12-13 [PMID: 24337741] (ICC, Immunoprecipitation, Western Blot, Human)
• Yan J, Zhang Y, Zhang J, Liang J, Li L, Zheng L Tim-3 expression defines regulatory T cells in human tumors. PLoS ONE, 2013-03-05;8(3):e58006. 2013-03-05 [PMID: 23526963] (IHC, Human)
• Dennie Tompers Frederick, Adriano Piris, Alexandria P. Cogdill, Zachary A. Cooper, Cecilia Lezcano, Cristina R. Ferrone et al. BRAF Inhibition Is Associated with Enhanced Melanoma Antigen Expression and a More Favorable Tumor Microenvironment in Patients with Metastatic Melanoma Clinical Cancer Research 3/1/2013 [PMID: 23307859]
• Dennie Tompers Frederick, Adriano Piris, Alexandria P. Cogdill, Zachary A. Cooper, Cecilia Lezcano, Cristina R. Ferrone et al. BRAF Inhibition Is Associated with Enhanced Melanoma Antigen Expression and a More Favorable Tumor Microenvironment in Patients with Metastatic Melanoma Clinical Cancer Research 2013-03-01 [PMID: 23307859] (Immunohistochemistry, Human)
• Zhang Y, Ma CJ, Wang JM, Ji XJ, Wu XY, Jia ZS, Moorman JP, Yao ZQ Tim-3 negatively regulates IL-12 expression by monocytes in HCV infection. PLoS ONE, 2011-05-26;6(5):e19664. 2011-05-26 [PMID: 21637332] (Neutralization, Human)
• Vega-Carrascal I, Reeves EP, Niki T, Arikawa T, McNally P, O'Neill SJ, Hirashima M, McElvaney NG Dysregulation of TIM-3-Galectin-9 Pathway in the Cystic Fibrosis Airways. J. Immunol., 2011-01-24;0(0):. 2011-01-24 [PMID: 21263071] (ICC, Immunoprecipitation, Western Blot, Human)
• Seki M, Sakata KM, Oomizu S, Arikawa T, Sakata A, Ueno M, Nobumoto A, Niki T, Saita N, Ito K, Dai SY, Katoh S, Nishi N, Tsukano M, Ishikawa K, Yamauchi A, Kuchroo V, Hirashima M Beneficial effect of galectin 9 on rheumatoid arthritis by induction of apoptosis of synovial fibroblasts. Arthritis Rheum., 2007-12-01;56(12):3968-76. 2007-12-01 [PMID: 18050192] (ICC, Neutralization, Human)

Reviews for TIM-3 Antibody (AF2365) (2)

Average Rating: 4.5

(Based on 2 reviews)

We have 2 reviews tested in 1 species: Human.

reviewed by: Verified Customer WB Human 04/04/2020

Summary

• Application: Western Blot
• Sample Tested: Jurkat
• Species: Human
• Lot: UAJ0218021

reviewed by: Verified Customer immunofluorescence - paraffin Human 05/02/2019

Summary

• Application: Other
• Sample Tested: human tonsil
• Species: Human
• Lot: N/A

Comments

• Comments: pH 9 heat-induced antigen retrieval

Bioinformatics

• Gene Symbol: HAVCR2
• Uniprot:
  • Q8TDQ0()