Cd3e Hamster Monoclonal Antibody [Clone ID: 145-2C11]

Specifications

Product Data
Clone Name	145-2C11
Applications	FC
Recommend Dilution	Flow Cytometry (See Protocols). This clone has also been reported to work in Immunoprecipitation 1,2 and Western Blotting 8.
Reactivity	Mouse
Host	Hamster
Clonality	Monoclonal
Immunogen	H-2Kb sp from Armenian Hamster Spleen. Fusion Partner: Murine myeloma cell line SP2/0
Specificity	This anti-Mouse CD3e monoclonal antibody is specific for a 25 kDa protein component (e-T3) of the antigen specific T cell receptor on all mouse strains tested. The e-T3 protein has been shown to be non-covalently associated on the cell surface ab heterodimer of the CD3 associated complex. This monoclonal antibody reacts with all mature T cells and can both activate and inhibit T cell function (1). This fact identifies e-T3 as a cell surface protein involved in the transduction of activation signals. All peripheral T cells express this determinant however B cells and bone marrow cells have proven to be negative. Although the expression of this particular epitope on peripheral T cells is uniformly high, staining of thymocytes reveals distinct subpopulations of cells differing in the level of expression of this marker. This antibody will prove useful in studying the role of various components of the TCR complex in T cell activation and development, and will allow for the development of an animal model in which to investigate the immunoregulatory effects of in vivo administration of anti-CD3 antibodies, an area of obvious clinical importance. Anti-CD3e is ideal for flow cytometry applications, particularly as a specific marker for tracking mouse T cells. In addition, this monoclonal antibody, clone 145-2C11, was specifically designed to trigger T cell activation.
Formulation	PBS, 0.2 µm sterile filtered Label: Biotin State: Liquid purified Ascites
Concentration	lot specific
Purification	Protein G Affinity Chromatography
Conjugation	Biotin
Storage Condition	Store undiluted at 2-8°C for one month or (in aliquots) at -20°C for longer. Avoid repeated freezing and thawing.
Gene Name	CD3 antigen, epsilon polypeptide
Database Link	Entrez Gene 12501 Mouse UniProt ID P22646
Background	T cell activation through the antigen receptor (TCR) involves the cytoplasmic tails of the CD3 subunits: CD3 gamma, CD3 delta, CD3 epsilon and CD3 zeta. These CD3 subunits are structurally related members of the immunoglobulins super family encoded by closely linked genes on human chromosome 11. The CD3 components have long cytoplasmic tails that associate with cytoplasmic signal transduction molecules. This association is mediated at least in part by a double tyrosine based motif present in a single copy in the CD3 subunits. CD3 may play a role in TCR induced growth arrest, cell survival and proliferation. The CD3 antigen is present on 68-82% of normal peripheral blood lymphocytes, 65-85% of thymocytes and Purkinje cells in the cerebellum. It is never expressed on B or NK cells. Decreased percentages of T lymphocytes may be observed in some autoimmune diseases.
Synonyms	T3/Leu-4
Note	Donor: BALB/c Cell Concentration: 1x106 cells Antibody Concentration: 1 µg/106 cells Isotypic Control: Biotin Hamster IgG Protocol: Flow Cytometry Analysis: 1. Prepare cell suspension in Media A. For cell preparations, deplete the red blood cell population with Lympholyte®-M cell separation medium. 2. Wash 2 times. 3. Resuspend cells to 1x106 cells in approximately 50 µl Media A in a microcentrifuge tube (i.e. 50 µl of cells resuspended to 2x107 cells/ml). (THE CONTENTS OF 1 TUBE REPRESENTS 1 TEST). 4. To each tube add 1 µg of this antibody per 106 cells*. 5. Vortex the tubes to ensure thorough mixing of antibody and cells. 6. Incubate the tubes for 30 minutes at 4°C. 7. Wash 2 times at 4°C. 8. Add 100 µl of secondary antibody at a 1/500 dilution. 9. Incubate tubes at 4°C for 30-60 minutes. (It is recommended that the tubes are protected from light since most fluorochromes are light sensitive). 10. Wash 2 times at 4°C in Media B. 11. Resuspend the cell pellet in 50 µl ice cold Media B. 12. Transfer to suitable tubes for flow cytometric analysis containing 15 µl of propidium iodide at 0.5 mg/ml in phosphate buffered saline. (This stains dead cells by intercalating DNA). Media: A. Phosphate buffered saline (pH 7.2) + 5% normal serum of host species + sodium azide ( 100 µl of 2 M sodium azide in 100 mls). B. Phosphate buffered saline (pH 7.2) + 0.5% bovine serum albumin + sodium azide (100 µl of 2 M sodium azide in 100 mls).
Reference Data

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