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Anti-NFAT1 antibody [25A10.D6.D2] (ab2722)

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Overview

Product name

Anti-NFAT1 antibody [25A10.D6.D2]
See all NFAT1 products (6) ...

Description

Mouse monoclonal [25A10.D6.D2] to NFAT1

Specificity

Ab2722 detects nuclear factor of activated T-cells (NFAT) from mouse, rat and human tissues (endogenously expressed). This antibody does not cross react with NFAT2 (NFATc, NFATc1). This antibody detects both forms NFAT1 - a ~140 kDa protein representing phosphorylated NFAT1 in resting immune cells, and a ~120 kDa protein in stimulated cells that represents fully-dephosphorylated NFAT1.

Tested applications

IHC-P, Gel supershift assays, ICC, IP, WBmore details

Cross reactivity

Reacts with

Mouse, Rat, Human

Immunogen

Synthetic peptide: (51)AISSPSGLAYPDDVLDYGL(69). The immunizing peptide corresponds to amino acid residues 51-69 of mouse NFAT1-A, B and C isoforms. The sequence of this N-terminal peptide is ~70% homologous with human NFAT1.

AISSPSGLAYPDDVLDYGL

Run BLAST with BLAST the sequence with ExPASyRun BLAST with BLAST the sequence with NCBI

Positive control

WB: resting immune cells and ionomycin stimulated immune cells (see Shaw et al reference: "1uM for T cells and B cells, 10uM for macrophages,and 0.3uM for mast cells. Treatment was for 20 min.")

Properties

Form

Liquid

Storage instructions

Store at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.

Storage buffer

PBS with 0.05% sodium azide

Concentration

Concentration information loading...

Purity

IgG fraction

Clonality

Monoclonal

Clone number

25A10.D6.D2

Isotype

IgG1

Research areas

Metabolism >> Types of disease >> Cancer
Metabolism >> Pathways and Processes >> Metabolism processes >> Hypoxia
Cancer >> Cancer Metabolism >> Response to hypoxia
Cardiovascular >> Heart >> Hypertrophy >> Transcription factors
Epigenetics and Nuclear Signaling >> Nuclear Signaling Pathways >> NFATs
Epigenetics and Nuclear Signaling >> Transcription >> Other factors
Signal Transduction >> Signaling Pathway >> Nuclear Signaling >> NFATS

Immunology >> Adaptive Immunity >> T Cells >> Non-CD

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - NFAT1 antibody [25A10.D6.D2] (ab2722)Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - NFAT1 antibody [25A10.D6.D2] (ab2722) image (enlarge)

  • Western blot - NFAT1 antibody [25A10.D6.D2] (ab2722)Western blot - NFAT1 antibody [25A10.D6.D2] (ab2722) image (enlarge)

  • Flow Cytometry-Anti-NFAT1 antibody [25A10.D6.D2](ab2722)Flow Cytometry-Anti-NFAT1 antibody [25A10.D6.D2](ab2722) image (enlarge)

Applications

Show applications key

Our Abpromise guarantee covers the use of ab2722 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

  • ShowHide1 Image

    IHC-P

     IHC-P: 1/125. PubMed: 19...Read more →

    IHC-P: 1/125. PubMed: 19828810

  • ShowHide

    Gel supershift assays

     GSA: Use at an assay dep...Read more →

    GSA: Use at an assay dependent dilution. PubMed: 12065418

  • ShowHide

    ICC

     ICC: Use a concentration...Read more →

    ICC: Use a concentration of 1 µg/ml. Fix with 3% paraformaldehyde at 37°C or with ice-cold methanol (see Shaw et al reference (clone refered to as 67.1).

  • ShowHide

    IP

     IP: Use at an assay depe...Read more →

    IP: Use at an assay dependent dilution.

  • ShowHide1 Image

    WB

    Three stars (2 Abreviews) WB: Use a concentration ...Read more →

    WB: Use a concentration of 1 µg/ml. Detects a band of approximately 120 , 140 (phospho) kDa (predicted molecular weight: 115 kDa).

Target

Function

Plays a role in the inducible expression of cytokine genes in T-cells, especially in the induction of the IL-2, IL-3, IL-4, TNF-alpha or GM-CSF.

Tissue specificity

Expressed in thymus, spleen, heart, testis, brain, placenta, muscle and pancreas.

Sequence similarities

Contains 1 RHD (Rel-like) domain.

Domain

Rel Similarity Domain (RSD) allows DNA-binding and cooperative interactions with AP1 factors.

Post-translational
modifications

In resting cells, phosphorylated by NFATC-kinase on at least 18 sites in the 99-363 region. Upon cell stimulation, all these sites except Ser-243 are dephosphorylated by calcineurin. Dephosphorylation induces a conformational change that simultaneously exposes an NLS and masks an NES, which results in nuclear localization. Simultaneously, Ser-53 or Ser-56 is phosphorylated; which is required for full transcriptional activity.

Cellular localization

Cytoplasm. Nucleus. Cytoplasmic for the phosphorylated form and nuclear after activation that is controlled by calcineurin-mediated dephosphorylation. Rapid nuclear exit of NFATC is thought to be one mechanism by which cells distinguish between sustained and transient calcium signals. The subcellular localization of NFATC plays a key role in the regulation of gene transcription.

Target information above from: UniProt accessionQ13469 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010).

Information by UniProt

Alternative names

  • cytoplasmic 2 antibody
  • AI607462 antibody
  • KIAA0611 antibody
  • NF ATp antibody
  • NF-ATc2 antibody
  • NF-ATp antibody
  • NFAC2_HUMAN antibody
  • NFAT 1 antibody
  • NFAT pre existing subunit antibody
  • NFAT pre-existing subunit antibody
  • NFAT transcription complex, preexisting component antibody
  • NFAT1 antibody
  • NFAT1-D antibody
  • NFATc2 antibody
  • NFATp antibody
  • Nuclear factor of activated T cells cytoplasmic 2 antibody
  • Nuclear factor of activated T cells cytoplasmic calcineurin dependent 2 antibody
  • Nuclear factor of activated T cells pre-existing component antibody
  • Nuclear factor of activated T-cells antibody
  • T cell transcription factor NFAT 1 antibody
  • T-cell transcription factor NFAT1 antibody
see all

Database links

Anti-NFAT1 antibody [25A10.D6.D2] images:

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - NFAT1 antibody [25A10.D6.D2] (ab2722)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - NFAT1 antibody [25A10.D6.D2] (ab2722)

ab2722 (4µg/ml) staining NFAT in human tonsil, using an automated system (DAKO Autostainer Plus). Using this protocol there is strong nuclear and weak cytoplasmic staining.
Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.

  Western blot - NFAT1 antibody [25A10.D6.D2] (ab2722)

Western blot - NFAT1 antibody [25A10.D6.D2] (ab2722)

Anti-NFAT1 antibody [25A10.D6.D2] (ab2722) at 1 µg/ml + Spleen (Human) Tissue Lysate - adult normal tissue (ab29699) at 10 µg

Secondary
Goat polyclonal Secondary Antibody to Mouse IgG - H&L (HRP), pre-adsorbed (ab97040) at 1/5000 dilution ( )
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 115 kDa
Observed band size : 150 kDa (why is the actual band size different from the predicted?)
Additional bands at : 62 kDa. We are unsure as to the identity of these extra bands.

Exposure time : 4 minutes

NFAT1 contains an exstensive number of potential phosphorylation sites (SwissProt) which may explain its migration at a higher molecular weight than predicted.

  Flow Cytometry-Anti-NFAT1 antibody [25A10.D6.D2](ab2722)

Flow Cytometry-Anti-NFAT1 antibody [25A10.D6.D2](ab2722)

Overlay histogram showing Jurkat cells stained with ab2722 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab2722, 2µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

References for Anti-NFAT1 antibody [25A10.D6.D2] (ab2722)

This product has been referenced in:

  • Szeto GL  et al. Minocycline Suppresses Activation of Nuclear Factor of Activated T Cells 1 (NFAT1) in Human CD4+ T Cells. J Biol Chem 286:11275-82 (2011). WB; Human.Read more (PubMed: 21282105) »
  • González-Navajas JM  et al. TLR4 signaling in effector CD4+ T cells regulates TCR activation and experimental colitis in mice. J Clin Invest 120:570-81 (2010). WB; Mouse.Read more (PubMed: 20051628) »

See all 8 publications for this product

Publishing research using ab2722? Please let us know so that we can cite the reference in this datasheet

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"