Recombinant Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)
![ChIP - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-9-AntiNanog-antibody-EPR20694--ChIP-GradeChIP.jpg "ChIP - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR20694] to Nanog - ChIP Grade
- Suitable for: Flow Cyt (Intra), ChIP, IHC-P, WB, ICC/IF, IP
- Reacts with: Mouse
Related conjugates and formulations
Overview
-
Product name
Anti-Nanog antibody [EPR20694] - ChIP Grade
See all Nanog primary antibodies -
Description
Rabbit monoclonal [EPR20694] to Nanog - ChIP Grade -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt (Intra), ChIP, IHC-P, WB, ICC/IF, IPmore details -
Species reactivity
Reacts with: Mouse -
Immunogen
Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.
-
Positive control
- WB: F9 and ES-D3 whole cell lysates. IHC-P: Mouse E14.5 testis tissue. ICC/IF: F9 cells. Flow Cyt (intra): F9 cells. IP: F9 whole cell lysate. ChIP: Chromatin prepared from F9 cells.
-
General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
-
Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR20694 -
Isotype
IgG -
Research areas
Associated products
-
Alternative Versions
-
Compatible Secondaries
-
Isotype control
-
Recombinant Protein
-
Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab214549 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 |
|---|---|---|
| Flow Cyt (Intra) |
1/60.
|
|
| ChIP |
Use 5 µg for 25 µg of chromatin.
|
|
| IHC-P |
1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
|
|
| WB |
1/1000. Detects a band of approximately 29-42 kDa (predicted molecular weight: 34 kDa).
|
|
| ICC/IF | (1) |
1/100.
|
| IP |
1/30.
|
| Notes |
|---|
|
Flow Cyt (Intra)
1/60. |
|
ChIP
Use 5 µg for 25 µg of chromatin. |
|
IHC-P
1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
|
WB
1/1000. Detects a band of approximately 29-42 kDa (predicted molecular weight: 34 kDa). |
|
ICC/IF
1/100. |
|
IP
1/30. |
Target
-
Function
Transcription regulator involved in inner cell mass and embryonic stem (ES) cells proliferation and self-renewal. Imposes pluripotency on ES cells and prevents their differentiation towards extraembryonic endoderm and trophectoderm lineages. Blocks bone morphogenetic protein-induced mesoderm differentiation of ES cells by physically interacting with SMAD1 and interfering with the recruitment of coactivators to the active SMAD transcriptional complexes (By similarity). Acts as a transcriptional activator or repressor (By similarity). Binds optimally to the DNA consensus sequence 5'-TAAT[GT][GT]-3' or 5'-[CG][GA][CG]C[GC]ATTAN[GC]-3' (By similarity). When overexpressed, promotes cells to enter into S phase and proliferation. -
Tissue specificity
Expressed in testicular carcinoma and derived germ cell tumors (at protein level). Expressed in fetal gonads, ovary and testis. Also expressed in ovary teratocarcinoma cell line and testicular embryonic carcinoma. Not expressed in many somatic organs and oocytes. -
Sequence similarities
Belongs to the Nanog homeobox family.
Contains 1 homeobox DNA-binding domain. -
Developmental stage
Expressed in embryonic stem (ES) and carcinoma (EC) cells. Expressed in inner cell mass (ICM) of the blastocyst and gonocytes between 14 and 19 weeks of gestation (at protein level). Not expressed in oocytes, unfertilized oocytes, 2-16 cell embryos and early morula (at protein level). Expressed in embryonic stem cells (ES). Expression decreases with ES differentiation. -
Cellular localization
Nucleus. - Information by UniProt
-
Database links
- Entrez Gene: 71950 Mouse
- SwissProt: Q80Z64 Mouse
- Unigene: 6047 Mouse
-
Alternative names
- Embryonic stem cell specific homeobox protein (Nanog) antibody
- ENK antibody
- FLJ12581 antibody
see all
Images
-
ChIP - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)
Chromatin was prepared from F9 cells according to the Abcam X-ChIP protocol. Cells were fixed with 1% formaldehyde for 10 minutes. The ChIP was performed with 25 µg of chromatin, 5µg of ab214549 (red), and 20 µl of protein A/G sepharose beads slurry (10 µl of sepharose A beads + 10 µl of sepharose G beads). Then 5 μg of rabbit normal IgG was added to the control beads (grey). The immunoprecipitated DNA was quantified by real time PCR (SYBR green chemistry).
-
![Western blot - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-300508-anti-nanog-antibody-epr20694-chip-grade-western-blot.jpg)
Western blot - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)All lanes : Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549) at 1/1000 dilution
Lane 1 : F9 (Mouse embryonic testicular cancer cell line) whole cell lysate
Lane 2 : NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Developed using the ECL technique.
Predicted band size: 34 kDa
Observed band size: 29-42 kDa why is the actual band size different from the predicted?
Exposure time: 70 secondsBlocking/Dilution buffer: 5% NFDM/TBST.
The multiple bands observed are consistent with the literature (PMID: 24936455). Negative control: NIH/3T3 (PMID: 12787505).
-
![Western blot - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-300509-anti-nanog-antibody-epr20694-chip-grade-western-blot.jpg)
Western blot - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549) at 1/1000 dilution + ES-D3 (mouse embryonic multipotent stem cell line) whole cell lysate at 10 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Developed using the ECL technique.
Predicted band size: 34 kDa
Observed band size: 29-42 kDa why is the actual band size different from the predicted?
Exposure time: 3 minutesBlocking/Dilution buffer: 5% NFDM/TBST.
The multiple bands observed are consistent with the literature (PMID: 24936455).
-
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-300511-anti-nanog-antibody-epr20694-chip-grade-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)Immunohistochemical analysis of paraffin-embedded mouse E14.5 testis tissue labeling Nanog with ab214549 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Mainly nuclear staining is observed in testis of mouse embryo E14.5 (PMID: 15939376; PMID: 12787505). Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
-
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-300512-anti-nanog-antibody-epr20694-chip-grade-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)Immunohistochemical analysis of paraffin-embedded adult mouse testis tissue labeling Nanog with ab214549 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Negative tissue: No staining on adult mouse testis is observed(PMID: 12787505; PMID: 15939376).
Counter stained with Hematoxylin.
Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
-
![Immunocytochemistry/ Immunofluorescence - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-300510-anti-nanog-antibody-epr20694-chip-grade-immunofluorescence.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeablized F9 (mouse embryonic testicular cancer cell line) and NIH/3T3 (mouse embryo fibroblast cell line) cells labeling Nanog with ab214549 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining in F9 cell line. Negative control: NIH/3T3 (PMID: 17352742).
The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.
Control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.
-
![Flow Cytometry (Intracellular) - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-300513-anti-nanog-antibody-epr20694-chip-grade-flow-cytometry.jpg)
Flow Cytometry (Intracellular) - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)Intracellular flow cytometric analysis of 4 % paraformaldehyde-fixed, 90% methanol permeabilized F9 (mouse embryonic testicular cancer cell line, Right) and NIH/3T3 (mouse embryo fibroblast cell line, Left) cells labeling Nanog with ab214549 at 1/60 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.
Negative control: NIH/3T3 cell line.
-
![Immunoprecipitation - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-300514-anti-nanog-antibody-epr20694-chip-grade-immunoprecipitation.jpg)
Immunoprecipitation - Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)Nanog was immunoprecipitated from 0.35 mg of F9 (mouse embryonic testicular cancer cell line) whole cell lysate with ab214549 at 1/1000 dilution. Western blot was performed from the immunoprecipitate using ab214549 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5,000 dilution
Lane 1: F9 whole cell lysate 10 μg (input)
Lane 2: ab214549 IP in F9 whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab214549 in F9 whole cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 15 seconds.
The multiple bands observed are consistent with the literature (PMID: 24936455).
-
![Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)](https://www.abcam.com/ps/products/214/ab214549/Images/ab214549-10-benefits-of-recombinant-antibodies.png)
Anti-Nanog antibody [EPR20694] - ChIP Grade (ab214549)
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
-
SDS download
-
Datasheet download
Certificate of Compliance
References (4)
ab214549 has been referenced in 4 publications.
- Tan A et al. TFEB regulates pluripotency transcriptional network in mouse embryonic stem cells independent of autophagy-lysosomal biogenesis. Cell Death Dis 12:343 (2021). PubMed: 33795648
- Li J et al. Aberrant Gcm1 expression mediates Wnt/ß-catenin pathway activation in folate deficiency involved in neural tube defects. Cell Death Dis 12:234 (2021). PubMed: 33664222
- Zhu Q et al. O-GlcNAcylation regulates the methionine cycle to promote pluripotency of stem cells. Proc Natl Acad Sci U S A 117:7755-7763 (2020). PubMed: 32193337
- Zhang S et al. The Hippo Effector Transcriptional Coactivator with PDZ-Binding Motif Cooperates with Oncogenic ß-Catenin to Induce Hepatoblastoma Development in Mice and Humans. Am J Pathol 190:1397-1413 (2020). PubMed: 32283103