The O4-46 monoclonal antibody specifically binds to human and mouse T-bet. T-bet (T-box gene expressed in T cells) is a master regulatory transcription factor that is also known as TBX21 (T-box21) and TBLYM (T-box transcription factor, expressed in lymphocytes). Human (535 amino acids; 58.3 kDa predicted molecular mass) and mouse (530 amino acids; 57.7 kDa) T-bet proteins are encoded by the human TBX21 (chromosome 17) and mouse Tbx21 (chromosome 11) genes. The human and mouse T-bet protein amino acid sequences are 88% homologous. Human and mouse T-bet proteins share a highly conserved (98% homologous amino acid sequences) T-box protein domain that is centrally located and mediates binding to DNA. T-bet is expressed by and activates transcriptional activities within hemotopoietic cells including stem cells, NK and NKT cells and subsets of thymocytes, primed/activated CD4+ T cells, CD8+ T cells and γδ T cells, B cells, and dendritic cells. Interferon-gamma (IFN-γ), interleukin-27 (IL-27), and IL-12 act on peripheral antigen-triggered (TCR-signaling) T cells to increase T-bet expression. With respect to T helper lymphocytes, T-bet directs the differentiation of naïve CD4+ precursor T cells to become Th1-like effector and memory cells. T-bet accomplishes this by activating Th1 genetic programs (including epigenetic modifications) while repressing opposing T helper subset programs. T-bet controls the upregulated expression of the Th1 signature cytokine, IFN-γ, the IL-12Rβ2 subunit and the Runx3 transcription factor and can repress the function of other transcriptional regulators, such as GATA-3 (master regulator of Th2 development) and the expression of other cytokines including IL-2, IL-4 and IL-5.
Immunofluorescent staining with the O4-46 Mouse Anti-T-bet monoclonal antibody is compatible with BD™ Phosflow Lyse/Fix Buffer (Cat. No. 558049) and the BD™ Phosflow Permeabilization Buffers: Perm/Wash Buffer I (Cat. No. 557885); Perm Buffer II (Cat. No. 558052); Perm Buffer III (Cat. No. 558050); and Perm Buffer IV (Cat. No. 560746).
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to Alexa Fluor® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed.
1. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
2. An isotype control should be used at the same concentration as the antibody of interest.
3. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
4. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
5. The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
6. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
7. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
8. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
9. This product may be covered by US Patent No. 7,365,169.
10. Limited Use License: The buyer (a) shall not sell or otherwise transfer this product to any third party, (b) shall use this product only for its internal, non-clinical research use, (c) shall not use this product for Commercial Purposes without a commercial license from Harvard (and if the buyer is interested in a commercial license, it should contact Harvard’s Office of Technology Development at 1350 Massachusetts Avenue, Holyoke Center, Suite 727, Cambridge, MA 02138, 617-495-3067), (d) shall use this product in compliance with all applicable laws and regulations, including, without limitation, applicable human health and animal welfare laws and regulations, (e) may transfer information or materials made through the use of this product to a scientific collaborator only if such transfer is not for any Commercial Purpose and such collaborator agrees in writing not to transfer such materials to any third party and to use such transferred materials and/or information solely for internal, non-clinical research and not for Commercial Purposes, (f) acknowledges that this product has not been approved for use in humans by the U.S. Food and Drug Administration or any other regulatory body and may not be used in humans and (g) shall indemnify, defend and hold harmless Becton Dickinson and Company and Harvard from and against all damages, losses, expenses (including reasonable attorneys’ fees), claims, demands, suits and other actions in any way arising from the buyer’s use, storage or disposal of this product.
11. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
12. Cy is a trademark of GE Healthcare.
13. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
