Recombinant Anti-TCF-4 antibody [NCI-R159-6] - BSA and Azide free (ab223073)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [NCI-R159-6] to TCF-4 - BSA and Azide free
- Suitable for: Flow Cyt (Intra), WB, IHC, ChIP
- Reacts with: Mouse, Human
Related conjugates and formulations
Overview
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Product name
Anti-TCF-4 antibody [NCI-R159-6] - BSA and Azide free
See all TCF-4 primary antibodies -
Description
Rabbit monoclonal [NCI-R159-6] to TCF-4 - BSA and Azide free -
Host species
Rabbit -
Specificity
Mouse specificity in WB only.Weak human specificity in IHC.
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Tested applications
Suitable for: Flow Cyt (Intra), WB, IHC, ChIPmore details -
Species reactivity
Reacts with: Mouse, Human -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: Nuclear extracts of Cal-1 cells infected with control TCF-4 shRNA. SH-SY5Y nuclear extracts. U-87 MG and Neuro-2a whole cell lysates, Daudi nuclear fraction, U-87 MG whole cell lysate, Neuro-2a whole cell lysate IHC: Human tonsil. Blastic plasmacytoid dendritic cell neoplasm (BPDCN) cell pellets after selection and induction of shRNA expression for 1 day. Flow Cyt (intra): Cal-1 cells. ChIP: Cal-1 cells
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General notes
ab223073 is the carrier-free version of ab217668.
For detailed protocol using this antibody for IHC, ChIP, and Flow Cyt, please refer to the following paper:
A Druggable TCF4- and BRD4-Dependent Transcriptional Network Sustains Malignancy in Blastic Plasmacytoid Dendritic Cell NeoplasmOur carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
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
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
pH: 7.2
Constituent: PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
NCI-R159-6 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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ChIP Related Products
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Conjugation kits
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Isotype control
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Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab223073 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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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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WB |
Use at an assay dependent concentration. Detects a band of approximately 90 kDa (predicted molecular weight: 71 kDa).
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IHC |
Use at an assay dependent concentration.
Fixative: 4% FormalinAntigen retrieval: Low pH retrieval solution. |
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ChIP |
Use at an assay dependent concentration.
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Notes |
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Flow Cyt (Intra)
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. Detects a band of approximately 90 kDa (predicted molecular weight: 71 kDa). |
IHC
Use at an assay dependent concentration. Fixative: 4% FormalinAntigen retrieval: Low pH retrieval solution. |
ChIP
Use at an assay dependent concentration. |
Target
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Function
Transcription factor that binds to the immunoglobulin enchancer Mu-E5/KE5-motif. Binds to the E-box present in the somatostatin receptor 2 initiator element (SSTR2-INR) to activate transcription (By similarity). Preferentially binds to either 5'-ACANNTGT-3' or 5'-CCANNTGG-3'. -
Tissue specificity
Expressed in adult heart, brain, placenta, skeletal muscle and to a lesser extent in the lung. In developing embryonic tissues, expression mostly occurs in the brain. -
Involvement in disease
Defects in TCF4 are a cause of Pitt-Hopkins syndrome (PTHS) [MIM:610954]. PTHS is a rare syndromic encephalopathy characterized by severe psychomotor delay, epilepsy, daily bouts of diurnal hyperventilation starting in infancy, mild postnatal growth retardation, postnatal microcephaly, and distinctive facial features. Since most hitherto reported cases have been sporadic, with males and females equally affected, PTHS is regarded as an autosomal dominant condition. -
Sequence similarities
Contains 1 basic helix-loop-helix (bHLH) domain. -
Domain
the 9aaTAD motif is a transactivation domain present in a large number of yeast and animal transcription factors. -
Cellular localization
Nucleus. - Information by UniProt
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Database links
- Entrez Gene: 6925 Human
- Entrez Gene: 21413 Mouse
- Omim: 602272 Human
- SwissProt: P15884 Human
- SwissProt: Q60722 Mouse
- Unigene: 605153 Human
- Unigene: 4269 Mouse
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Alternative names
- bHLHb19 antibody
- Class B basic helix-loop-helix protein 19 antibody
- E2 2 antibody
see all
Images
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All lanes : Anti-TCF-4 antibody [NCI-R159-6] (ab217668) at 1/10000 dilution
Lane 1 : Nuclear extracts of Cal-1 cells (Human plasmacytoid dendritic cell line) infected with control TCF4 shRNA
Lane 2 : Nuclear extracts of Cal-1 cells infected with TCF4 shRNA
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated
Predicted band size: 71 kDa
Observed band size: 90 kDa why is the actual band size different from the predicted?
Exposure time: 40 secondsThe data was provided by the collaborator Dr. Louis M. Staudt, NCI, NIH.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
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All lanes : Anti-TCF-4 antibody [NCI-R159-6] (ab217668) at 1/1000 dilution
Lane 1 : Daudi (human Burkitt's lymphoma lymphoblast) cytoplasmic fraction
Lane 2 : Daudi nuclear fraction
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.
Performed under reducing conditions.
Predicted band size: 71 kDa
Observed band size: 58,79 kDa why is the actual band size different from the predicted?
Exposure time: 3 minutesExposure time : 3 minutes
Blocking/Dilution buffer: 5% NFDM/TBST.
This blot was developed using a high sensitivity ECL substrate.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
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Immunohistochemical analysis of 4% Formalin fixed Blastic plasmacytoid dendritic cell neoplasm (BPDCN) cell pellets after selection and induction of shRNA expression for 1 day, labeling TCF-4 with ab217668 at 1/100 dilution. Universal DAB Detection Kit was used for detection of IHC staining on an automated system.
The data was provided by our collaborator Dr. Louis M. Staudt, and published in Cancer Cell 30, 764-778, 2016 (PMID: 27846392). Several TCF-4 shRNAs were used. This IHC image shows data for shRNA TCF-4 #2.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
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All lanes : Anti-TCF-4 antibody [NCI-R159-6] (ab217668) at 1/1000 dilution
Lane 1 : U-87 MG (human glioblastoma-astrocytoma epithelial cell), whole cell lysate
Lane 2 : Neuro-2a (mouse neuroblastoma neuroblast), 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.
Performed under reducing conditions.
Predicted band size: 71 kDa
Observed band size: 58,79 kDa why is the actual band size different from the predicted?
Exposure time: 3 minutesExposure time : 3 minutes
Blocking/Dilution buffer: 5% NFDM/TBST.
This blot was developed using a high sensitivity ECL substrate.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
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Cal-1 cells (Human plasmacytoid dendritic cell line) were cross-linked with 1% formaldehyde for 5 min at RT. Cross-linked cells were first washed with ice-cold PBS and then resuspended in ice-cold RIPA buffer (10mM Tris-HCl pH8, 140 mM NaCl, 1mM EDTA pH 8, 0.5 mM EGTA, 1% Triton X-100, 0.1% SDS and 0.1% Sodium Deoxycholate) to a final concentration of 5x106 cells/ml. DNA was sheared with a Misonix XL sonicator, by performing 12 x 45’’ sonication cycles at power setting of 5. For each ChIP reaction, 2x107 chromatin cell equivalents were incubated overnight with10 μg of ab217668. The following day, chromatin/antibody complexes were incubated with 50 μl of Protein G/Protein A magnetic beads mix (G to A ratio 3:1) for 4 h at 4°C. Normal rabbit IgG was added to the beads as control.
The TCF-4 locus ChIP-seq tracks for BRD4 (blue), RNA Pol2 (red), and TCF-4 (green) are shown for Cal-1 cells. This data was kindly provided by our collaborator Dr. Louis M. Staudt, and has been published (PMID: 27846392).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
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Intracellular flow cytometric analysis of 1% paraformaldehyde-fixed, ice-cold methanol permeabilized Cal-1 cells (Human plasmacytoid dendritic cell line) (black - positive control) and Cal-1 cells infected with either Ctrl (left green) or TCF4 (right green) shRNA, labeling TCF4 with ab217668 at 1/100 dilution (green and black) compared with a Rabbit IgG control (grey). Goat anti-Rabbit IgG (Alexa Fluor® 647) at 1/500 dilution was used as the secondary antibody.
The data was provided by our collaborator Dr. Louis M. Staudt, and published in Cancer Cell 30, 764-778, 2016 (PMID: 27846392). Several TCF4 shRNAs were used. This FC image shows data for shRNA TCF4
1.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
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Immunohistochemical analysis of 4% Formalin fixed human tonsil labeling TCF-4 with ab217668 at 1/100 dilution. Universal DAB Detection Kit was used for detection of IHC staining on an automated system.
pDCs: plasmacytoid dendritic cells.
GC: Germinal Center.
The data was provided by our collaborator Dr. Louis M. Staudt, and published in Cancer Cell 30, 764-778, 2016 (PMID: 27846392).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
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Anti-TCF-4 antibody [NCI-R159-6] (ab217668) at 1/200 dilution + SH-SY5Y (Human neuroblastoma cell line from bone marrow) nuclear extracts at 10 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 71 kDa
Observed band size: 90 kDa why is the actual band size different from the predicted?This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab217668).
Blocking and Diluting buffer and concentration: 5% NFDM /TBST
The isoforms expression pattern is consistent with the literatures (PMID: 21789225).
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
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Datasheet download
Certificate of Compliance
References (1)
ab223073 has been referenced in 1 publication.
- Shi X et al. EWS-FLI1 regulates and cooperates with core regulatory circuitry in Ewing sarcoma. Nucleic Acids Res 48:11434-11451 (2020). PubMed: 33080033