Anti-NFAT5 antibody (ab3446)
Key features and details
- Rabbit polyclonal to NFAT5
- Suitable for: IHC-P, ICC/IF, WB, IP
- Reacts with: Mouse, Human, African green monkey
- Isotype: IgG
Overview
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Product name
Anti-NFAT5 antibody
See all NFAT5 primary antibodies -
Description
Rabbit polyclonal to NFAT5 -
Host species
Rabbit -
Specificity
Detects Nuclear Factor of Activated T-cells 5 (NFAT 5). -
Tested applications
Suitable for: IHC-P, ICC/IF, WB, IPmore details -
Species reactivity
Reacts with: Mouse, Human, African green monkey -
Immunogen
Synthetic peptide corresponding to Human NFAT5 aa 1439-1455 (C terminal).
Sequence:DLLVSLQNQGNNLTGSF
(Peptide available asab4978) -
Positive control
- WB: MCF7, Jurkat, Raji, Ramos, HepG2, U2OS, HeLa, COS7, EL4, C2C12 and NRK cell lysates. IHC-P: Human brain and skeletal muscle tissues. ICC/IF: NIH/3T3, MCF7 and HeLa cells. IP: U2OS cells.
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General notes
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
Properties
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Form
Liquid -
Storage instructions
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. -
Storage buffer
Preservative: 0.05% Sodium azide
Constituents: 0.1% BSA, 99% PBS -
Concentration information loading...
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Purity
Immunogen affinity purified -
Clonality
Polyclonal -
Isotype
IgG -
Research areas
Associated products
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ChIP Related Products
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Compatible Secondaries
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Immunizing Peptide (Blocking)
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Isotype control
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Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab3446 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 |
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IHC-P |
1/20. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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ICC/IF |
1/100.
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WB | (1) |
1/1000. Detects a band of approximately 170 kDa (predicted molecular weight: 160 kDa).Can be blocked with NFAT5 peptide (ab4978).
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IP |
Use at an assay dependent concentration.
3 μg |
Notes |
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IHC-P
1/20. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
ICC/IF
1/100. |
WB
1/1000. Detects a band of approximately 170 kDa (predicted molecular weight: 160 kDa).Can be blocked with NFAT5 peptide (ab4978). |
IP
Use at an assay dependent concentration. 3 μg |
Target
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Function
Plays a role in the inducible expression of genes. Regulates hypertonicity-induced cellular accumulation of osmolytes. -
Tissue specificity
Highest levels in skeletal muscle, brain, heart and peripheral blood leukocytes. Also expressed in placenta, lung, liver, kidney, pancreas, spleen, thymus, prostate, testis, ovary, small intestine and colon. -
Sequence similarities
Contains 1 RHD (Rel-like) domain. -
Cellular localization
Nucleus. - Information by UniProt
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Database links
- Entrez Gene: 10725 Human
- Entrez Gene: 54446 Mouse
- Omim: 604708 Human
- SwissProt: O94916 Human
- SwissProt: Q9WV30 Mouse
- Unigene: 371987 Human
- Unigene: 390057 Mouse
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Alternative names
- Glutamine rich protein H65 antibody
- KIAA0827 antibody
- NF AT5 antibody
see all
Images
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All lanes : Anti-NFAT5 antibody (ab3446) at 1/1000 dilution
Lane 1 : MCF7 (Human breast adenocarcinoma cell line) whole cell lysate
Lane 2 : Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate
Lane 3 : Raji (Human Burkitt's lymphoma cell line) whole cell lysate
Lane 4 : Ramos (Human Burkitt's lymphoma cell line) whole cell lysate
Lane 5 : HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate
Lane 6 : U-2 OS (Human bone osteosarcoma epithelial cell line) whole cell lysate
Lane 7 : HeLa (Human epithelial adenocarcinoma cell line) whole cell lysate
Lane 8 : COS-7 (African green monkey kidney fibroblast-like cell line) whole cell lysate
Lane 9 : EL4 (Mouse thymic lymphoma cell line) whole cell lysate
Lane 10 : C2C12 (Mouse myoblast cell line) whole cell lysate
Lane 11 : NRK (Rat kidney normal tissue) whole cell lysate
Lysates/proteins at 25 µg per lane.
Secondary
All lanes : Goat anti-rabbit-HRP secondary antibody at 1/20000 dilution
Predicted band size: 160 kDaWestern blot analysis of NFAT5 was performed by loading samples onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% Milk/TBST for at least 1 hour. Membranes were incubated with ab3446 overnight at 4°C on a rocking platform. Membranes were washed in TBS-0.1%Tween 20 and probed with a secondary antibody for at least one hour. Membranes were washed and chemiluminescent detection performed.
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ICC analysis of NFAT5 in HeLa (Human epithelial adenocarcinoma cell line) cells. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control - right) or with ab3446 at a dilution of 1:20 overnight at 4 C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. NFAT5 staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Images were taken at 60X magnification.
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Immunohistochemistry was performed on normal biopsies of deparaffinized human skeletal muscle tissue. 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 1/20 duution with ab3446 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.
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Immunoprecipitation of NFAT5 was performed on U-2 OS (Human bone osteosarcoma epithelial cell line) whole cell lysate (lane 2). The antigen:antibody complex was formed by incubating 500 µg whole cell lysate with 3 µg of ab3446 overnight on a rocking platform at 4°C. The immune-complex was captured on 50 µL Protein A/G Plus Agarose. Captured immune-complexes were washed and proteins eluted with 5X Reducing Sample Loading Dye. Samples were resolved on a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to PVDF membrane and blocked with 5% Milk/TBS-0.1%Tween for at least 1 hour. Membranes were washed in TBS-0.1%Tween 20 and probed with a goat anti-rabbit-HRP secondary antibody at a dilution of 1/20,000 for at least one hour. Membranes were washed and chemiluminescent detection performed.
Lane 1: Only cell lysate.
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ICC analysis of NFAT5 in NIH/3T3 (Mouse embryo fibroblast cell line) cells. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control - right) or with ab3446 at a dilution of 1:20 overnight at 4 C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. NFAT5 staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Images were taken at 60X magnification.
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Immunohistochemistry was performed on normal biopsies of deparaffinized human brain tissue. 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 1:20 with ab3446 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.
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ICC analysis of NFAT5 in MCF7 (Human breast adenocarcinoma cell line) cells. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control - right) or with ab3446 at a dilution of 1:200 overnight at 4 C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. NFAT5 staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Images were taken at 60X magnification.
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ICC analysis of NFAT5 usingab3446 (shown in green) in HeLa (Human epithelial adenocarcinoma cell line) whole cells. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature. Cells were then blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with a rabbit polyclonal antibody recognizing NFAT5, at a dilution of 1/100 for at least 1 hour at room temperature. Cells were washed with PBS and incubated with DyLight 488 goat-anti-rabbit secondary antibody at a dilution of 1/400 for 30 minutes at room temperature. Nuclei (blue) were stained with Hoechst 33342 dye. Images were taken on a Thermo Scientific ArrayScan at 20X magnification.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (31)
ab3446 has been referenced in 31 publications.
- Zhang H et al. Long noncoding RNA KCNQ1OT1 inhibits osteoclast differentiation by regulating the miR-128-3p/NFAT5 axis. Aging (Albany NY) 14:4486-4499 (2022). PubMed: 35587369
- Chernyakov D et al. The nuclear factor of activated T cells 5 (NFAT5) contributes to the renal corticomedullary differences in gene expression. Sci Rep 12:20304 (2022). PubMed: 36433977
- Duan YR et al. LncRNA lnc-ISG20 promotes renal fibrosis in diabetic nephropathy by inducing AKT phosphorylation through miR-486-5p/NFAT5. J Cell Mol Med 25:4922-4937 (2021). PubMed: 33939247
- Dou Y et al. Circ_0001944 Contributes to Glycolysis and Tumor Growth by Upregulating NFAT5 Through Acting as a Decoy for miR-142-5p in Non-Small Cell Lung Cancer. Cancer Manag Res 13:3775-3787 (2021). PubMed: 34040437
- Chernyakov D et al. Loss of RANBP3L leads to transformation of renal epithelial cells towards a renal clear cell carcinoma like phenotype. J Exp Clin Cancer Res 40:226 (2021). PubMed: 34233711