Biolegend, 285463, Spark NIR™ 685 anti-mouse CD3ε (Flexi-Fluor™) Antibody, 50μg

CD3ε is a 20 kD transmembrane protein, also known as CD3 or T3. It is a member of the Ig superfamily and primarily expressed on T cells, NK-T cells, and at different levels on thymocytes during T cell differentiation. CD3ε forms a TCR complex by associating with the CD3δ, γ and ζ chains, as well as the TCR α/β or γ/δ chains. CD3 plays a critical role in TCR signal transduction, T cell activation, and antigen recognition by binding the peptide/MHC antigen complex.
50μg
Verified Reactivity: Mouse
Antibody Type: Monoclonal
Host Species: Armenian Hamster
Immunogen: H-2Kb-specific mouse cytotoxic T lymphocyte clone BM10-37
Formulation: Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide
Preparation: The antibody was purified by affinity chromatography and conjugated with Spark NIR™ 685 under optimal conditions.
Concentration: 0.2 mg/mL
Storage & Handling: The antibody solution should be stored undiluted between 2°C and 8°C, and protected from prolonged exposure to light. Do not freeze.
Application: FC
Recommended Usage: Flexi-Fluors™ are provided at a standard 0.2 mg/mL concentration. We recommend titrating this reagent to determine the optimal concentration for each application. For many flow cytometry applications, conjugated antibodies perform well at concentrations ranging from 0.03 to 1.0 µg per million cells in 100 µL. We recommend testing a range of concentrations starting from 10 µg/mL. For example, make five 1:1 serial dilutions of the 0.2 mg/mL antibody. Add 5 µL of each dilution (including the undiluted antibody) to 100 µL of cells (at 107 cells/mL) to test six concentrations -- 1.0, 0.5, 0.25, 0.125, 0.06, and 0.03 µg per million cells in 100 µL volume. Compare staining patterns or create a titration curve using the MFI or staining index to determine the optimal concentration. * Spark NIR™ 685 has a maximum excitation of 665 nm and a maximum emission of 685 nm.
Excitation Laser: Red Laser (633 nm)
Application Notes: Clone 145-2C11 is useful for in vitro blocking of target-specific CTL-mediated cell lysis1, as well as T cell activation assays, inducing proliferation and cytokine production1,2,7,12,16. It also induces apoptosis in immature thymocytes32, and in vivo T cell depletion8-10. Additional reported applications (for relevant formats of this clone) include: immunoprecipitation1, immunohistochemical staining14,15 of acetone-fixed frozen sections and zinc-fixed paraffin-embedded sections, Western blotting4, complement-mediated cytotoxicity6, in vitro and in vivo stimulation of T cells1,2,7,12,16, immunofluorescent staining5, and in vivo T cell depletion8-10. The 145-2C11 antibody has been reported to block the binding of 17A2 antibody to CD3 epsilon-specific T cells11. Clone 145-2C11 is not recommended for formalin-fixed paraffin embedded sections. The LEAF™ purified antibody (Endotoxin <0.1 EU/µg, Azide-Free, 0.2 µm filtered) is recommended for functional assays (Cat. No. 100314). For in vivo studies or highly sensitive assays, we recommend Ultra-LEAF™ purified antibody (Cat. No. 100340) with a lower endotoxin limit than standard LEAF™ purified antibodies (Endotoxin <0.01 EU/µg).
Additional Product Notes: For more information about Flexi-Fluors™, visit our Flexi-Fluor™ page and review FAQs associated with this product line.
Application References(PubMed link indicates BioLegend citation): Leo O, et al. 1987. P. Natl. Acad. Sci. USA 84:1374. (IP, Activ, Block) Kruisbeek AM, et al. 1991. In Current Protocols in Immunology. 3.12.1. (Activ) Duke RC, et al. 1995. Current Protocols in Immunology. 3.17.1. Salvadori S, et al. 1994. J. Immunol. 153:5176. (WB) Payer E, et al. 1991. J. Immunol. 146:2536. (IF) Jacobs H, et al. 1994. Eur. J. Immunol. 24:934. (CMCD) Vossen ACTM, et al. 1995. Eur. J. Immunol. 25:1492. (Activ) Henrickson M, et al. 1995. Transplantation 60:828. (Deplete) Kinnaert P, et al. 1996. Transpl. Int. 9:386. (Deplete) Han WR, et al. 1999. Transpl. Immunol. 7:207. (Deplete) Miescher GC, et al. 1989. Immunol. Lett. 23:113. (Block) Terrazas LI, et al. 2005. Intl. J. Parasitology. 35:1349. (Activ) Ko SY, et al. 2005. J. Immunol. 175:3309. Podd BS, et al. 2006. J. Immunol. 176:6532. (IHC-F) Tilley SL, et al. 2007. J. Immunol. 178:3208. (IHC-F) Wang W, et al. 2007. J. Immunol. 178:4885. (Activ) Xiao S, et al. 2007. J. Exp. Med. 204:1691. Chappaz S, et al. 2007. Blood doi:10.1182/blood-2007-02-074245. (FC) PubMed. Curtsinger JM, et al.2005. J. Immunol. 175:4392. PubMed Guo Y, et al. 2008. Blood 112:480. PubMed Kenna TJ, et al. 2008. Blood 111:2091. Perchonock CE, et al. 2007. J. Immunol. 179:1768. PubMed Perchonock GE, et al. 2006. Mol. Cell. Biol. 26:6005. PubMed Kanaya T, et al. 2008. Am. J. Physiol. Gastrointest. Liver Physiol. 295:G273. PubMed de Koning BA, et al. 2006. Int. Immunol. 18:941. PubMed Schulteis RD, et al. 2008. Blood 295:G273. PubMed Qi Q, et al. 2009. Blood 114:564. PubMed Helmersson S, et al. 2013. Am J Pathol. 9440:123. Pubmed Wu S, et al. 2014. Clin Vaccine Immunol. 21:156. PubMed Yan J, et al. 2014. Vaccine. 32:2833. PubMed Guiterrez DA, et al. 2014. Diaebetes. 63:3827. PubMed Shi YF, et al. 1991. J Immunol. 146:3340. (Apop)
RRID: AB_3716946 (BioLegend Cat. No. 285463)
Structure: Ig superfamily, forms CD3/TCR complex with CD3δ, γ and ζ subunits and TCR (α/β and γ/δ), 20 kD
Distribution: Thymocytes (differentiation dependent), mature T cells, NK-T cells
Function: TCR signal transduction, T cell activation, antigen recognition
Ligand/Receptor: Peptide antigen/MHC-complex
Cell Type: NKT cells, T cells, Thymocytes, Tregs
Biology Area: Immunology
Molecular Family: CD Molecules, TCRs
Antigen References: 1. Barclay A, et al. 1997. The Leukocyte Antigen FactsBook Academic Press. 2. Davis MM. 1990. Annu. Rev. Biochem. 59:475. 3. Weiss A, et al. 1994. Cell 76:263.
Gene ID: 12501
UniProt: View information about CD3epsilon on UniProt.org
Clone: 145-2C11
Regulatory Status: RUO
Other Names: CD3ε, T3, CD3
Isotype: Armenian Hamster IgG
Q: What are Flexi-Fluors?
A: Flexi-Fluors are rapidly made-to-order conjugated antibodies. The technology, manufacturing processes, and specifications used to create Flexi-Fluors are the same as our regular catalog products. However, the optimal concentration and performance of each Flexi-Fluor must be determined by the customer.
Q: How quickly will I receive my order?
A: We aim to ship Flexi-Fluors within 2-3 weeks of receipt of your order. However, depending on your location, shipping times may vary.
Q: How are Flexi-Fluors different from regular catalog products?
A: Flexi-Fluors are made on demand, specifically for you. Flexi-Fluors are manufactured using the same high-quality standards, and specifications as other catalog products. For faster delivery, Flexi-Fluors are not tested by flow cytometry to determine optimal concentrations or evaluate performance. This testing needs to be performed by the customer.
Q: How do I determine the optimal concentration for using my Flexi-Fluor? How should I titrate my antibody?
A: Flexi-Fluors are provided at a standard 0.2 mg/mL concentration. We recommend that you titrate your antibody to determine the optimal concentration to use for your application. For many flow cytometry applications, conjugated antibodies perform well at concentrations ranging from 0.03 to 1.0 µg per million cells in 100 µL volume. We recommend that you test a range of concentrations starting from 10 µg/mL. 

For example, make five 1:1 serial dilutions of your 0.2 mg/mL antibody.  Add 5 µL of each dilution (including the undiluted antibody) to 100 µL of cells (at 107 cells/ml) to test six concentrations - 1.0, 0.5, 0.25, 0.125, 0.06, and 0.03 µg per million cells in 100 µL volume. Compare staining patterns or create a titration curve using the MFI or staining index to determine the optimal concentration.
Q: I can’t find the antibody-dye combination that I need. When will it be available?
A: We continuously update our catalog, introducing scores of new products every month. Please get in touch with our Technical Service team for an update on new products or recommendations for suitable alternatives to complete your panel. Or contact Custom Solutions to inquire about our affordable custom conjugation services.
Q: I need help to validate the performance of my Flexi-Fluor. Who should I contact?
A: Please get in touch with Technical Service for assistance.
Q: Can I order more than 50 μg of a Flexi-Fluor?
A: Yes, you can order multiple vials of the same Flexi-Fluor products.  We cannot guarantee, however, that these vials will be bottled from the same lot. For bulk single-lot orders, contact our Custom Solutions team.
Q: What is the expiration date of my Flexi-Fluor?
A: Expiration dates can be found on the vial label or by using our CoA lookup tool.
Q: What if my requested antibody-dye combination doesn't perform as expected?
A: Flexi-Fluor™ products have been carefully selected by our scientists for their excellent conjugation properties and broad compatibility. In the rare case that an antibody-dye combination does not perform as expected due to the unique characteristics of a specific pairing, please contact our Technical Service team. We will work with you to identify alternative options to meet your panel requirements.