Overview

  • Product name

    Anti-NFAT4/NF-ATc3 (phospho S165) antibody
    See all NFAT4/NF-ATc3 primary antibodies
  • Description

    Rabbit polyclonal to NFAT4/NF-ATc3 (phospho S165)
  • Host species

    Rabbit
  • Specificity

    Detects endogenous levels of NFAT4/NF-ATc3 only when phosphorylated at serine 165.

  • Tested applications

    Suitable for: ICC/IF, WB, ELISAmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to Human NFAT4/NF-ATc3. (Around the phosphoryation site of serine 165 (SPSPPA))

  • General notes

    Previously labelled as NFAT4.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at -20°C. Stable for 12 months at -20°C.
  • Storage buffer

    pH: 7.40
    Preservative: 0.02% Sodium azide
    Constituents: PBS, 50% Glycerol, 0.87% Sodium chloride

    Without Mg+2 and Ca+2
  • Concentration information loading...
  • Purity

    Immunogen affinity purified
  • Purification notes

    Affinity purified from rabbit antiserum by affinity chromatography using epitope specific phosphopeptide. The antibody against non-phosphopeptide was removed by chromatography using non-phosphopeptide corresponding to the phosphorylation site.
  • Clonality

    Polyclonal
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab59204 in the following tested applications.

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

Application Abreviews Notes
ICC/IF Use at an assay dependent concentration. PubMed: 21889978
WB 1/500 - 1/1000. Detects a band of approximately 116 kDa (predicted molecular weight: 116 kDa).
ELISA 1/5000.

Target

  • Function

    Plays a role in the inducible expression of cytokine genes in T-cells, especially in the induction of the IL-2.
  • Tissue specificity

    Isoform 1 is predominantly expressed in thymus and is also found in peripheral blood leukocytes and kidney. Isoform 2 is predominantly expressed in skeletal muscle and is also found in thymus, kidney, testis, spleen, prostate, ovary, small intestine, heart, placenta and pancreas. Isoform 3 is expressed in thymus and kidney. Isoform 4 is expressed in thymus and skeletal muscle.
  • 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

    Phosphorylated by NFATC-kinase; dephosphorylated by calcineurin.
  • 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.
  • Information by UniProt
  • Database links

  • Alternative names

    • C80703 antibody
    • cytoplasmic 3 antibody
    • D8Ertd281e antibody
    • NF AT4 antibody
    • NF ATc3 antibody
    • NF-AT4 antibody
    • NF-ATc3 antibody
    • NFAC3_HUMAN antibody
    • NFAT4 antibody
    • NFATc3 antibody
    • NFATx antibody
    • Nuclear factor of activated T cells, cytoplasmic 3 antibody
    • nuclear factor of activated T cells, cytoplasmic, calcineurin dependent 3 antibody
    • Nuclear factor of activated T-cells antibody
    • T cell transcription factor NFAT4 antibody
    • T-cell transcription factor NFAT4 antibody
    see all

Images

  • All lanes : Anti-NFAT4/NF-ATc3 (phospho S165) antibody (ab59204) at 1/500 dilution

    Lane 1 : HeLa cells treated with calcium (40nM, 30 mins)
    Lane 2 : HeLa cells treated with calcium (40nM, 30 mins) with immunizing phosphopeptide

    Predicted band size: 116 kDa
    Observed band size: 116 kDa

References

This product has been referenced in:

  • Saygili E  et al. Rate and irregularity of electrical activation during atrial fibrillation affect myocardial NGF expression via different signalling routes. Cell Signal : (2011). WB, ICC/IF ; Rat . Read more (PubMed: 21889978) »
  • Lin Z  et al. miR-23a functions downstream of NFATc3 to regulate cardiac hypertrophy. Proc Natl Acad Sci U S A 106:12103-8 (2009). WB ; Rat . Read more (PubMed: 19574461) »
See all 2 Publications for this product

Customer reviews and Q&As

Question
Answer



The cell treatment protocol is as follows:

a. HeLa cells were plated on flasks or dishes containing growth medium (basic medium containing 10% serum).
b. When the cells were at the log phase and 80-90% confluent, the culture media was drained. Wash cells with fresh basic media (no serum) once.
c. Add fresh basic media to the vessels and starve the cells for 18-24 hours.
d. Drain the cultured medium, then add fresh basic medium (about 4ml per T75 flask) to the vessels.
e. Add Ca2+ to the medium at 40nM.

f. Cells were incubated at 37°C for 30 mins in a CO2 incubator and then were used to prepare cell lysates.



Cell lysates were prepared as follows:
a. For adherent cells, wash cells with Wash Buffer (4℃) once. Add 5ml Wash Buffer (4℃) per T75 flask and immediately scrape the cells off with a rubber scraper. Collect cells by centrifugation (3000rpm, 5min) at 4℃ and remove Wash Buffer.
b. Leave the tube containing cells on ice for 10 minutes. Then add Cold (4℃) Lysis Buffer to the cells (150ul per 1X106-1X107cells).
c. Mix the cells in the buffer and roll tube continuous on ice for 30 minutes at 4℃.
d. (Optional) Sonication for 4X8 at output level 5 for whole cell lysis.
e. Collect the supernatant by centrifugation (>12000rhm, 10minutes) at 4℃. Assay protein concentration by BCA. The cell lysis can be stored at -70℃ or ready to use.

Wash buffer formula: 10 mM PBS with 1 mM NaF and 1 mM Na3VO4. Lysis buffer formula: 50 mM Kcl with 1% NP40, 25 mM HEPES, 125 uM DTT, and protease/phosphatase inhibitors (100 ug/mL leupeptin, 20 ug/mL aprotinin, PMSF 1 mM, Na3VO4 1 mM).

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