A20/TNFAIP3 Antibody (59A426) - Azide and BSA Free

Novus Biologicals | Catalog # NBP2-33252

Novus Biologicals
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Key Product Details

Species Reactivity

Human, Mouse, Rat

Applications

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, Flow Cytometry, Flow (Intracellular), Immunocytochemistry/ Immunofluorescence, Simple Western, Immunoprecipitation

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 kappa Clone # 59A426

Format

Azide and BSA Free
View all A20/TNFAIP3 Primary Antibodies »

Product Specifications

Immunogen

Full length recombinant human A20/TNFAIP3 Antibody (59A426). The epitope has been mapped to the C-terminal portion of A20 (amino acids 440-790)

Clonality

Monoclonal

Host

Mouse

Isotype

IgG1 kappa

Scientific Data Images for A20/TNFAIP3 Antibody (59A426) - Azide and BSA Free

Western Blot: A20/TNFAIP3 Antibody (59A426)Azide Free [NBP2-33252]

Western Blot: A20/TNFAIP3 Antibody (59A426)Azide Free [NBP2-33252]

Western Blot: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252] - Human Jurkat lysate probed with A20 antibody at 4 ug/ml. Image from the standard format of this antibody.
Immunocytochemistry/ Immunofluorescence: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Immunocytochemistry/ Immunofluorescence: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Immunocytochemistry/Immunofluorescence: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252] - HepG2 cells were fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X PBS + 0.05% Triton-X100. The cells were incubated with anti-A20/TNFAIP3 (59A426) at 10 ug/ml overnight at 4C and detected with an anti-mouse Dylight 488 (Green) at a 1:500 dilution. Actin was detected with Phalloidin 568 (Red) at a 1:200 dilution. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective Image from the standard format of this antibody.
Immunohistochemistry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Immunohistochemistry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Immunohistochemistry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252] - Human placenta probed with A20 antibody at 5 ug/ml, cytoplasmic staining of decidual cells is observed. Human tissue TMA was used for this test. Staining of formalin-fixed tissues is enhanced by boiling tissue sections in 10 mM sodium citrate buffer, pH 6
Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252] - Intracellular flow cytometric analysis of A20 in ThP-1 cells using A20 antibody (red) and isotype control (green) at 0.5 ug/ml Image from the standard format of this antibody.
Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252] - An intracellular stain was performed on RH-30 cells with A20/TNFAIP3 Antibody (59A426) NBP1-77533PE (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeablized with 0.1% saponin. Cells were incubated in an antibody dilution of 2.5 ug/mL for 30 minutes at room temperature. Both antibodies were directly conjugated to Phycoerythrin.
Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252]

Flow Cytometry: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252] - An intracellular stain was performed on SK-MEL-28 cells with A20/TNFAIP3 (59A426) antibody NBP1-77533AF647 (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeablized with 0.1% saponin. Cells were incubated in an antibody dilution of 2.5 ug/mL for 30 minutes at room temperature. Both antibodies were directly conjugated to Alexa Fluor 647.
Simple Western: A20/TNFAIP3 Antibody (59A426)Azide Free [NBP2-33252]

Simple Western: A20/TNFAIP3 Antibody (59A426)Azide Free [NBP2-33252]

Simple Western: A20/TNFAIP3 Antibody (59A426) - Azide Free [NBP2-33252] - Lane view shows a specific band for A20/TNFAIP3 in 0.05 mg/ml of Jurkat lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system. Image from the standard format of this antibody.

Applications for A20/TNFAIP3 Antibody (59A426) - Azide and BSA Free

Application
Recommended Usage

Flow (Intracellular)

reported in scientific literature (Hjelmeland (2010))

Flow Cytometry

1ul/1 million cells

Immunocytochemistry/ Immunofluorescence

1:10-1:500

Immunohistochemistry-Paraffin

1:10-1:500. Use reported in scientific literature (Metellus (2010))

Immunoprecipitation

1-2 ug/ml

Western Blot

2-4 ug/ml
Application Notes
Multiple A20 cleavage fragments have been described in Western Blot, see Coornaert (2008) and Hailfinger (2009) for additional details.

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Flow Cytometry

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

Azide and BSA Free

Preservative

No Preservative

Concentration

0.5 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at -20C. Avoid freeze-thaw cycles.

Background: A20/TNFAIP3

A20, TNFalpha-induced protein 3, TNFAIP3 (theoretical molecular weight 95 kDa) is an important regulator of pro-inflammatory signaling pathways such as NF-kB activation and tumor necrosis factor (TNF)-mediated programmed cell death (1, 2). In macrophages, A20/TNFAIP3 controls the release of IL-1 beta/IL-18 by regulating NLRP3 inflammasome activity and CXCL9/CXCL10 production via STAT1 signaling. By regulating the expression of inflammatory molecules such as IL-6 and anti-apoptotic proteins in dendritic cells, A20/TNFAIP3 maintains T-cell and B-cell homeostasis.

A20/TNFAIP3 is a highly conserved protein sharing >90% amino acid sequence identity across various mammalian species and is highly expressed in T-cell and B-cells. Coding and non-coding single nucleotide polymorphisms (SNPs) of A20/TNFAIP3 have been associated with multiple autoinflammatory and autoimmune diseases including type 1 diabetes, rheumatoid arthritis, Crohn's disease, and systemic lupus erythematosus (SLE) (3). The two domains of A20/TNFAIP3 cooperate to regulate NF-kB signaling. Its N-terminal ovarian tumor (OTU) domain contains a catalytic cysteine (C103) which functions as a K63 deubiquitinase, whereas the 7 zinc fingers that make up the C-terminal domain mediate K48 polyubiquitination. To regulate NF-kB signaling, A20/TNFAIP3 removes K63-polyubiquitin chains from receptor-interacting protein 1 (RIP1) and NF-kB essential modulator (NEMO), thus preventing interactions with downstream partners. A20/TNFAIP3 also contributes to the degradation of RIP1 and Ubc13 through the addition of K48 polyubiquitin chains (4).

References

1.Dixit VM1, Green S, Sarma V, Holzman LB, Wolf FW, O'Rourke K, Ward PA, Prochownik EV, Marks RM. (1990) Tumor necrosis factor-alpha induction of novel gene products in human endothelial cells including a macrophage-specific chemotaxin. J Biol Chem. 265(5):2973-8. PMID: 2406243

2.Verstrepen L, Verhelst K, van Loo G, Carpentier I, Ley SC, Beyaert R. (2010) Expression, biological activities and mechanisms of action of A20 (TNFAIP3). Biochem Pharmacol. 80(12):2009-20. PMID: 20599425

3.Mele A, Cervantes JR, Chien V, Friedman D, Ferran C. (2014) Single nucleotide polymorphisms at the TNFAIP3/A20 locus and susceptibility/resistance to inflammatory and autoimmune diseases. Adv Exp Med Biol. 809:163-83. PMID: 25302371

4.Das T, Chen Z, Hendriks RW, Kool M. (2018) A20/Tumor Necrosis Factor alpha-Induced Protein 3 in Immune Cells Controls Development of Autoinflammation and Autoimmunity: Lessons from Mouse Models. Front Immunol. 9:104. PMID: 29515565

Alternate Names

OTUD7C, TNFA1P2, TNFAIP3

Gene Symbol

TNFAIP3

Additional A20/TNFAIP3 Products

Product Documents for A20/TNFAIP3 Antibody (59A426) - Azide and BSA Free

Certificate of Analysis

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Product Specific Notices for A20/TNFAIP3 Antibody (59A426) - Azide and BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

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Protocols

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FAQs for A20/TNFAIP3 Antibody (59A426) - Azide and BSA Free

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  • Q: Following cleavage of A20/TNFAIP3, two full length fragments are released, one with 37 kDa and other one was approx. at 52 kDa. There are mentioning of 75Kda fragment and 37 KDa fragment in scientific papers but not 52 KDa. What is the significance of that specific band?

    A: There is data available that explains 52 KDa band in A20/TNFAIP3. Using this antibody in Western Blot will result multiple bands. In LPS treatment, 52 KDa putative cleavage band was observed; however, the same band was not found in untreated THP1 cells.

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