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

• 产品名称

  Anti-NFAT1抗体[25A10.D6.D2]
  参阅全部 NFAT1 一抗

• 描述

  小鼠单克隆抗体[25A10.D6.D2] to NFAT1

• 宿主

  Mouse

• 特异性

  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.

• 经测试应用

  适用于: Flow Cyt, WB, IHC-P, ICC/IFmore details

• 种属反应性

  与反应: Human
  

• 免疫原

  Synthetic peptide corresponding to Mouse NFAT1 aa 50-150.
  Database link: Q60591

  Run BLAST with Run BLAST with 

• 阳性对照

• 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.")

• 常规说明

  
  

  The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

  If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

• 形式

  Liquid

• 存放说明

  Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.

• 存储溶液

  Preservative: 0.05% Sodium azide
  Constituent: PBS

• 纯度

  Protein G purified

• 克隆

  单克隆

• 克隆编号

  25A10.D6.D2

• 同种型

  IgG1

The Abpromise guarantee

Abpromise™承诺保证使用ab2722于以下的经测试应用

“应用说明”部分 下显示的仅为推荐的起始稀释度；实际最佳的稀释度/浓度应由使用者检定。

• 数据库链接

• Entrez Gene: 4773 Human

• Omim: 600490 Human

• SwissProt: Q13469 Human

• Unigene: 713650 Human

• 别名

• NF ATc2 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, preexisting component antibody

• Nuclear factor of activated T-cells antibody

• Preexisting nuclear factor of activated T cells 2 antibody

• T cell transcription factor NFAT 1 antibody

• T-cell transcription factor NFAT1 antibody

• AI607462 antibody

• cytoplasmic 2 antibody

• KIAA0611 antibody

• 

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

  Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human spleen tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a mouse monoclonal antibody recognizing NFATc2 ab2722 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.
  
  

• 

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

  Immunocytochemistry/Immunofluorescence analysis of NFAT1 (green) in HeLa cells. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% BSA for 15 minutes at room temperature. Cells were left untreated (left panel) or treated with 1uM staurosporine (right panel) for 3 hours and incubated with ab2722 (1:100) for at least 1 hour at room temperature, washed with PBS, and incubated with a DyLight 488 conjugated goat anti-mouse IgG secondary antibody (1:400) for 30 minutes at room temperature. F-Actin (red) was stained with DyLight 554 Phalloidin and nuclei (blue) were stained with Hoechst 33342 dye. Images were taken at 20X magnification.

• 

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

  Anti-NFAT1 antibody [25A10.D6.D2] (ab2722) at 1 µg/ml + Human spleen tissue lysate - total protein (ab29699) at 10 µg
  
  Secondary
  Goat Anti-Mouse IgG H&L (HRP) preadsorbed (ab97040) at 1/5000 dilution ( )
  
  Developed using the ECL technique.
  
  Performed under reducing conditions.
  
  Predicted band size: 115 kDa
  Observed band size: 150 kDawhy 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)

  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.
  
  

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-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.
  

• 

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

  Immunocytochemistry/Immunofluorescence analysis of NFAT1 (green) shows staining in HeLa cells. F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were inbuated without (control) or with ab2722 (1:20) over night at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

• 

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

  Immunocytochemistry/Immunofluorescence analysis of NFAT1 (green) shows staining in MCF-7 cells. F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were inbuated without (control) or with ab2722 (1:20) over night at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

• 

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

  Immunocytochemistry/Immunofluorescence analysis of NFAT1 (green) shows staining in U251 Cells. F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were inbuated without (control) or with ab2722 (1:20) over night at 4°C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Images were taken at 60X magnification.

• 

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

  Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human colon carcinoma tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a mouse monoclonal antibody recognizing NFATc2 ab2722 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.
  
  

• 

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

  Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized Human tonsil tissue tissues. To expose target proteins heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1:100 with a mouse monoclonal antibody recognizing NFATc2 ab2722 or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.