Explore
Validate
Learn
Western blot
ImmunoprecipitationAntibody data
- Antibody Data
- Antigen structure
- References [4]
- Comments [0]
- Validations
- Western blot [3]
- Immunocytochemistry [1]
- Chromatin Immunoprecipitation [1]
Submit
Validation data
Reference
Comment
Report error
- Product number
- 14-5824-82 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- T-bet Monoclonal Antibody (eBio39D (39D, 3-9D)), eBioscience™
- Antibody type
- Monoclonal
- Antigen
- Other
- Description
- Description: The eBio39D monoclonal antibody reacts with mouse, rat and human T-bet. T-bet is a Th1-specific T-box transcription factor critical to the development of the Th1 CD4+ T cell lineage. This is known based on the observations that T-Bet deficient mice have impaired Th1 cell development, and that ectopic expression of T-Bet results in development skewed to the Th1 lineage. T-Bet expression is induced by the Th1 cytokine IFN-gamma, and T-Bet also regulates the expression of IFN-gamma, likely, at least in part, through the modification of DNA accessibility and histone remodeling. In addition to IFN-gamma, T-Bet is also known to regulate the expression of IL-12Rbeta and IL-2. Moreover, T-Bet plays a role in class-switch recombination in B-cells. Applications Reported: This eBio39D (39D, 3-9D) antibody has been reported for use in immunoprecipitation, and immunoblotting (WB). Applications Tested: This eBio39D (39D, 3-9D) antibody has been tested by western blot analysis of Th1-polarized mouse CD4+ T cells and can be used at 2 µg/mL as a starting dilution for western blotting. Purity: Greater than 90%, as determined by SDS-PAGE. Aggregation: Less than 10%, as determined by HPLC. Filtration: 0.2 µm post-manufacturing filtered.
- Reactivity
- Human, Mouse, Rat
- Host
- Mouse
- Isotype
- IgG
- Antibody clone number
- eBio39D (39D, 3-9D)
- Vial size
- 100 µg
- Concentration
- 0.5 mg/mL
- Storage
- 4° C
Submitted references AMBRA1 Controls Regulatory T-Cell Differentiation and Homeostasis Upstream of the FOXO3-FOXP3 Axis.
T-bet regulates Th1 responses through essential effects on GATA-3 function rather than on IFNG gene acetylation and transcription.
IL-2 production in developing Th1 cells is regulated by heterodimerization of RelA and T-bet and requires T-bet serine residue 508.
A novel transcription factor, T-bet, directs Th1 lineage commitment.
Becher J, Simula L, Volpe E, Procaccini C, La Rocca C, D'Acunzo P, Cianfanelli V, Strappazzon F, Caruana I, Nazio F, Weber G, Gigantino V, Botti G, Ciccosanti F, Borsellino G, Campello S, Mandolesi G, De Bardi M, Fimia GM, D'Amelio M, Ruffini F, Furlan R, Centonze D, Martino G, Braghetta P, Chrisam M, Bonaldo P, Matarese G, Locatelli F, Battistini L, Cecconi F
Developmental cell 2018 Dec 3;47(5):592-607.e6
Developmental cell 2018 Dec 3;47(5):592-607.e6
T-bet regulates Th1 responses through essential effects on GATA-3 function rather than on IFNG gene acetylation and transcription.
Usui T, Preiss JC, Kanno Y, Yao ZJ, Bream JH, O'Shea JJ, Strober W
The Journal of experimental medicine 2006 Mar 20;203(3):755-66
The Journal of experimental medicine 2006 Mar 20;203(3):755-66
IL-2 production in developing Th1 cells is regulated by heterodimerization of RelA and T-bet and requires T-bet serine residue 508.
Hwang ES, Hong JH, Glimcher LH
The Journal of experimental medicine 2005 Nov 7;202(9):1289-300
The Journal of experimental medicine 2005 Nov 7;202(9):1289-300
A novel transcription factor, T-bet, directs Th1 lineage commitment.
Szabo SJ, Kim ST, Costa GL, Zhang X, Fathman CG, Glimcher LH
Cell 2000 Mar 17;100(6):655-69
Cell 2000 Mar 17;100(6):655-69
No comments: Submit comment
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- CD4+ T cells were sorted from mouse splenocytes, then activated with Anti-Mouse CD3 and Anti-Mouse CD28, followed by culture in Th1-polarizing conditions, and re-stimulation with PMA and Ionomycin. Lysates from control (left) or PMA and Ionomycin-re-stimulated (right) cells were probed with Anti-T-bet Purified at 2 µg/mL, and revealed with Anti-Mouse IgG HRP.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of T-bet was achieved by transfecting NK-92 with T-bet specific siRNAs (Silencer® select Product # s26889, s223839). Western blot analysis (Fig. a) was performed using modified whole cell extracts (1% SDS) from the T-bet knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with T-bet Monoclonal Antibody (Product # 14-5824-82, 2ug/ml) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 0.25µg/ml, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to T-bet.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-T-bet Monoclonal Antibody (Product # 14-5824-82) and a 58kDa band corresponding to T-bet was observed across cell lines tested and the protein was not expressed in K-562. Modified whole cell extracts (1% SDS) (30 µg lysate) of NK-92 (Lane 1) and K-562 (Lane 2) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (2ug/ml) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of T-bet was performed using 70% confluent log phase NK-92 cells. The cells were fixed with 4% paraformaldehyde for 10 Minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with T-bet Monoclonal Antibody (Product # 14-5824-82) at 5 µg/mL in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Mouse IgG (H+L), Superclonal™ Recombinant Secondary Antibody, Alexa Fluor 488 (Product # A28175) at a dilution of 1:2,000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415). Panel d represents the merged image showing nuclear localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Chromatin Immunoprecipitation (ChIP) assay of endogenous T-bet protein using Anti-T-bet Antibody: ChIP was performed using Anti-T-bet Mouse Monoclonal Antibody (Product # 14-5824-82, 5 µg) on sheared chromatin from NK-92 cells using the MAGnify ChIP System kit (Product # 49-2024). Normal Mouse IgG was used as a negative IP control. The purified DNA was analyzed by qPCR using primers binding to IFNG promoter, IKZF3 and CD19 transcriptional start site and SAT2 satellite repeats. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.
