Product Description
This Human TIM-3 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of TIM-3 protein (Cat: 10390-H03H) from the overexpression lysate was verified.
Expression Host
HEK293 Cells
Sequence Information
A DNA sequence encoding the human TIMD3 (NP_116171.3) extracellular domain (Met 1-Arg 200) was fused with the C-terminal polyhistidine-tagged Fc region of human IgG1 at the C-terminus.
Molecule Mass
The recombinant human TIMD3/Fc is a disulfide-linked homodimer after removal of the signal peptide. The reduced monomer consists of 426 amino acids and has a predicted molecular mass of 47.7 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rhTIMD3/Fc monomer is approximately 66 kDa due to glycosylation.
Preparation Method
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
Sample Buffer
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.
Human CD366 Overexpression Lysate;Human HAVcr-2 Overexpression Lysate;Human HAVCR2 Overexpression Lysate;Human KIM-3 Overexpression Lysate;Human Tim-3 Overexpression Lysate;Human TIM3 Overexpression Lysate;Human TIMD-3 Overexpression Lysate;Human TIMD3 Overexpression Lysate
Hepatitis A virus cellular receptor 2 (HAVCR2), formerly known as T cell immunoglobulin and mucin domain-3 (TIM-3), is a transmembrane glycoprotein expressed on the surface of terminally differentiated Th1 cells but not on Th2 cells. It was the first surface molecule that specifically identifies Th1 cells in both mice and human. Recently, identification of Galectin-9 as a ligand for TIM-3 has established the TIM-3-Galectin-9 pathway as an important regulator of Th1 immunity and tolerance induction. Engagement of Tim-3 by its ligand galectin-9 negatively regulates IFN-gamma secretion and influences the ability to induce T cell tolerance in both mice and man. It suggests a novel paradigm in which dysregulation of the TIM-3-galectin-9 pathway could underlie chronic autoimmune disease states, such as multiple sclerosis. Recent work has explored the role of TIM-3 in systemic lupus erythematosus (SLE), and their results indicate that TIM-3 may represent a novel target for the treatment of SLE. Numerous studies have demonstrated that Tim-3 influences autoimmune diseases, including diabetes and multiple sclerosis, and its role in other inflammatory diseases including allergies and cancer is beginning to become clear. In tumor rejection model, soluble form of Tim-3 (sTim-3) significantly impaired T cell antitumor immunity, evidenced by decreased antitumor CTL activity and reduced amount of tumor-infiltrating lymphocytes in tumor. sTim-3 as an immunoregulatory molecule that may be involved in the negative regulation of T cell-mediated immune response.
Immune Checkpoint Immune Checkpoint Detection: ELISA Antibodies Immune Checkpoint Detection: IP Antibodies Immune Checkpoint Detection: WB Antibodies Immune Checkpoint Proteins Immune Checkpoint Targets Co-inhibitory Immune Checkpoint Targets Immunotherapy Cancer Immunotherapy Targeted Therapy References
Geng H, et al. (2006) Soluble form of T cell Ig mucin 3 is an inhibitory molecule in T cell-mediated immune response. J Immunol. 176(3): 1411-20. Anderson AC, et al. (2006) TIM-3 in autoimmunity. Curr Opin Immunol. 18(6): 665-9. Anderson DE. (2007) TIM-3 as a therapeutic target in human inflammatory diseases. Expert Opin Ther Targets. 11(8): 1005-9. Pan HF, et al. (2010) TIM-3 as a new therapeutic target in systemic lupus erythematosus. Mol Biol Rep. 37(1): 395-8. 
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