Biolegend, 320127, True-Nuclear™ One Step Human Treg Flow Kit, 25tests

FOXP3 is a 50-55 kD transcription factor, also known as Forkhead box protein P3, Scurfin, JM2, or IPEX. It is proposed to be a master regulatory gene and more specific marker of T regulatory cells than most cell surface markers (such as CD4 and CD25). Transduced expression of FOXP3 in CD4 + /CD25 - cells has been shown to induce GITR, CD103, and CTLA4 and impart a T regulatory cell phenotype. FOXP3 is mutated in X-linked autoimmunity-allergic dysregulation syndrome (XLAAD or IPEX) in humans and in "scurfy" mice. Overexpression of FOXP3 has been shown to lead to a hypoactive immune state suggesting that this transcriptional factor is a central regulator of T cell activity. In human, unlike in mouse, two isoforms of FOXP3 have been reported: one (FOXP3) corresponding to the canonical full-length sequence; the other (FOXP3 δ2) lacking exon 2. The 206D antibody recognizes human FOXP3 epitope in the region of amino acids 105-235.
25tests
Verified Reactivity: Human
Reported Reactivity: Baboon, Cynomolgus, Rhesus, Pigtailed Macaque
Antibody Type: Monoclonal
Host Species: Mouse
Immunogen: Full-length FOXP3 protein
Concentration: Lot-specific (to obtain lot-specific concentration and expiration, please enter the lot number in our Certificate of Analysis online tool.)
Storage & Handling: This kit is guaranteed for six months. Upon receipt, store between 2°C and 8°C, and protected from prolonged exposure to light. Do not freeze.
Application: ICFC - Quality tested
Recommended Usage: Materials Provided: 1. Alexa Fluor® 488 anti-human FOXP3/CD25 PE/CD4 PerCP antibody cocktail - 25 tests 2. Alexa Fluor® 488 Mouse IgG1, κ isotype control/CD25 PE/CD4 PerCP antibody cocktail - 25 tests 3. True-Nuclear™ Transcription Buffer Set - 120 tests (Cat. No. 424401) Materials not included: 1. Cell Staining Buffer (Cat. No. 420201) 2. Single Color Compensation Controls Immunofluorescence Staining Procedures: 1. Aliquot 100 µL of target cells to each tube. 2. Add 1 mL of the Transcription Factor 1X Fix solution to each tube, vortex, and incubate at room temperature in the dark for 30-60 minutes. 3. Without washing, add 2 mL of the Transcription Factor 1X Perm Buffer to each tube. 4. Centifuge tubes at 400 x g at room temperature for five minutes, and discard the supernatant. 5. Add 2 mL of the Transcription Factor 1X Perm Buffer to each tube. 6. Centrifuge tubes at 400 x g at room temperature for five minutes, and discard the supernatant. 7. Resuspend the cell pellet in 100 µL of the Transcription Factor 1X Perm Buffer. 8. Add 20 µL of Alexa Fluor® 488 anti-human FOXP3/CD25 PE/CD4 PerCP antibody cocktail or 20 µL of Alexa Fluor® 488 mouse IgG1, κ isotype control/CD25 PE/CD4 PerCP antibody cocktail into the appropriate tubes. Incubate in the dark at room temperature for at least 30 minutes. 9. Add 2 mL of the Transcription Factor 1X Perm Buffer to each tube. 10. Centrifuge tubes at 400 x g at room temperature for five minutes, and discard the supernatant. 11. Add 2 mL of the cell staining buffer. 12. Centrifuge tubes at 400 x g at room temperature for five minutes, and discard the supernatant. 13. Resuspend in 0.5 mL cell staining buffer and then acquire tubes on a flow cytometer. Caution: The True-Nuclear™ Transcription Factor Buffer Set contains paraformaldehyde, which is toxic and mutagenic. Please handle with caution. Wear gloves, lab coats, and necessary protection to avoid direct contact. NOTE: For flow cytometric staining with this clone, True-Nuclear™ Transcription Factor Buffer Set (Cat. No. 424401) offers improved staining and is highly recommended over the Foxp3/Perm Buffer Set (Cat. No. 421403) and the Nuclear Factor Fixation and Permeabilization Buffer Set (Cat. No. 422601).
Application Notes: Additional reported applications (for the relevant formats) include: immunohistochemical staining of acetone-fixed frozen sections1 and formalin-fixed paraffin-embedded sections1, 8, 19-20, and Western blotting1. The binding of 206D to FOXP3 can be partially blocked by 259D, but 206D does not show significant blocking effect on 259D binding. NOTE: For flow cytometric staining with this clone, True-Nuclear™ Transcription Factor Buffer Set (Cat. No. 424401) offers improved staining and is highly recommended.
Application References(PubMed link indicates BioLegend citation): Roncador G, et al. 2005. Eur. J. Immunol. 35:1681. (IHC) Yang ZZ, et al. 2006. Blood 107:3639. Liu W, et al. 2006. J. Exp. Med. 203:1701.PubMed Bollyky PL, et al. 2007. J. Immunol. 179:744. Bell MP, et al. 2007. J. Immunol. 179:1893. Tran DQ, et al. 2007. Blood doi:10.1182/blood-2007-06-094656. PubMed Gao Q,et al.2007.J Clin Oncol.25:2586.(IHC) PubMed Pillai V,et al. 2008. Blood 111:463.PubMed Zheng Y, et al. 2008. J. Immunol. 181:1683. PubMed Zonios DI, et al. 2008. Blood112:287. PubMed Kavanagh B, et al. 2008. Blood. PubMed Nevala WK, et al. 2009. Clin Cancer Res. 15:1931. PubMed Grant J, et al. 2009. Cytometry B Clin Cytom. 76:69. PubMed Nigam P, et al. 2010. J. Immunol. 184:1690. PubMed Kmieciak M, et al. 2009. J. Transl. Med. 7:89. (ICFC) PubMed Hartigan-O'Connor DJ,et al. 2007. J Exp Med.204:2679. PubMed Raghaven S, et al. 2009. Ann Rheum Dis. 68:1908. PubMed Hodi FS, et al. 2014. Cancer Immunol Res. 2:632.(IHC) PubMed Sziros E, et al. 2015. Clin Cancer Res. 21:2840.(IHC) PubMed
Product Citations: Park S, et al. 2016. Proc Natl Acad Sci U S A. 113(52):E8379-E8386. PubMed
RRID: AB_3097006 (BioLegend Cat. No. 320127)
Structure: Forkhead/winged-helix transcription factor family, approximately 50 kD, contains zinc finger and forkhead domains
Distribution: Nuclear; expressed in T regulatory cells
Function: Transcription factor proposed to be a master regulatory gene in T regulatory cell development and a critical factor for immune homeostasis
Interaction: Interacts with DNA
Cell Type: Tregs
Biology Area: Cell Biology, Immunology, Transcription Factors
Molecular Family: CD Molecules, Nuclear Markers
Antigen References: 1. Hori S, et al. 2003. Science 299:1057. 2. Gandhi R, et al. 2010. Nat. Immunol. 11:846.
Regulation: FOXP3 is present at high levels in T regulatory cells, it can also be induced by T cell activation.
Gene ID: 509435094320371317382916
UniProt: View information about FOXP3 on UniProt.org
Clone: 206D
Regulatory Status: RUO
Other Names: Forkhead box protein P3, Scurfin, JM2, IPEX, Zinc finger protein JM2
Isotype: Mouse IgG1, κ
Q: Can I stain whole blood with anti-FOXP3 using your Foxp3 staining kit?
A: It is not recommended. It is best to use PBMCs for this testing.