Recombinant Anti-LEF1 antibody [EPR2029Y] - BSA and Azide free (ab215999)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR2029Y] to LEF1 - BSA and Azide free
- Suitable for: IP, ICC/IF, IHC-P, WB, Flow Cyt (Intra)
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-LEF1 antibody [EPR2029Y] - BSA and Azide free
See all LEF1 primary antibodies -
Description
Rabbit monoclonal [EPR2029Y] to LEF1 - BSA and Azide free -
Host species
Rabbit -
Tested applications
Suitable for: IP, ICC/IF, IHC-P, WB, Flow Cyt (Intra)more details -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: Jurkat cell lysate. IHC-P: Human tonsil tissue. Flow cyto (intra): Jurkat cells
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General notes
ab215999 is the carrier-free version of ab137872.
Our 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
EPR2029Y -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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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 ab215999 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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IP |
Use at an assay dependent concentration.
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ICC/IF |
Use at an assay dependent concentration.
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 44 kDa.
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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Notes |
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IP
Use at an assay dependent concentration. |
ICC/IF
Use at an assay dependent concentration. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
WB
Use at an assay dependent concentration. Predicted molecular weight: 44 kDa. |
Flow Cyt (Intra)
Use at an assay dependent concentration. |
Target
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Function
Participates in the Wnt signaling pathway. Activates transcription of target genes in the presence of CTNNB1 and EP300. May play a role in hair cell differentiation and follicle morphogenesis. TLE1, TLE2, TLE3 and TLE4 repress transactivation mediated by LEF1 and CTNNB1. Regulates T-cell receptor alpha enhancer function. Binds DNA in a sequence-specific manner. PIAG antagonizes both Wnt-dependent and Wnt-independent activation by LEF1 (By similarity). Isoform 3 lacks the CTNNB1 interaction domain and may be an antagonist for Wnt signaling. Isoform 5 transcriptionally activates the fibronectin promoter, binds to and represses transcription from the E-cadherin promoter in a CTNNB1-independent manner, and is involved in reducing cellular aggregation and increasing cell migration of pancreatic cancer cells. Isoform 1 transcriptionally activates MYC and CCND1 expression and enhances proliferation of pancreatic tumor cells. -
Tissue specificity
Detected in thymus. Not detected in normal colon, but highly expressed in colon cancer biopsies and colon cancer cell lines. Expressed in several pancreatic tumors and weakly expressed in normal pancreatic tissue. Isoforms 1 and 5 are detected in several pancreatic cell lines. -
Sequence similarities
Belongs to the TCF/LEF family.
Contains 1 HMG box DNA-binding domain. -
Domain
Proline-rich and acidic regions are implicated in the activation functions of RNA polymerase II transcription factors. -
Cellular localization
Nucleus. Found in nuclear bodies upon PIASG binding. - Information by UniProt
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Database links
- Entrez Gene: 51176 Human
- Entrez Gene: 16842 Mouse
- Entrez Gene: 161452 Rat
- Omim: 153245 Human
- SwissProt: Q9UJU2 Human
- SwissProt: P27782 Mouse
- SwissProt: Q9QXN1 Rat
- Unigene: 726506 Human
see all -
Alternative names
- DKFZp586H0919 antibody
- FLJ46390 antibody
- LEF 1 antibody
see all
Images
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
Flow cytometry overlay histogram showing left Jurkat positive cells and right negative HeLa stained with ab137872 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab137872) (1x 106 in 100μl at 0.2μg/ml (1/11500)) for 30min at 22°C.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C
Isotype control antibody (black line) was Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.
Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.
This antibody gave a positive signal in Jurkat Fixed with 4% formaldehyde (10 min) / permeabilised with 0.1% PBS-Triton X-100 for 15 min under the same conditions.
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All lanes : Anti-LEF1 antibody [EPR2029Y] (ab137872) at 1/1000 dilution
Lane 1 : Wild-type Jurkat cell lysate at 40 µg
Lane 2 : Lef1 knockout Jurkat cell lysate at 40 µg
Lane 3 : Jurkat cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 44 kDa
Observed band size: 40 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-LEF1 antibody [EPR2029Y] staining at 1/1000 dilution, shown in black; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab137872 was shown to bind specifically to LEF1. A band was observed at 40/53 kDa in wild-type Jurkat cell lysates with no signal observed at this size in Lef1 knockout cell line ab274898 (knockout cell lysate ab274956). To generate this image, wild-type and Lef1 knockout Jurkat cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times before development with Optiblot (ECL reagent ab133456) and imaged with 4 minutes exposure time. Secondary antibodies used were HRP conjugated Goat anti-Rabbit (H+L) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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Immunofluorescence staining of Jurkat (Human T cell leukemia cell line from peripheral blood) cells with purified ab137872 at a working dilution of 1 in 500, counter-stained with DAPI. Tubulin was stained with mouse anti-tubulin at a dilution of 1/1000 (ab7291) and Alexa Fluor® 594 goat anti-mouse at a dilution of 1/500 (ab150120) . The secondary antibody was ab150077 Alexa Fluor® 488 goat anti rabbit, used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100.
The negative controls are shown in the bottom middle and right hand panels - for the first negative control, purified ab137872 was used at a dilution of 1/200 followed by an Alexa Fluor® 555 goat anti-mouse antibody at a dilution of 1/500 and for the second negative control mouse primary antibody (ab7291) and anti-rabbit secondary antibody (ab15007) were used.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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Lane 1 (input): Rat thymus lysate, 10μg
Lane 2 (+): Rat thymus lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab137872 in rat thymus lysateab137872 immunoprecipitating LEF1 in rat thymus lysate. For western blotting, primary antibody used was ab137872 at 1:1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.
Blocking and diluting buffer: 5% NFDM/TBST
Exposure time: 3 minutes
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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Lane 1 (input): Jurkat (human T cell leukemia T lymphocyte) whole cell lysate, 10 μg
Lane 2 (+): Jurkat whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab137872 in Jurkat whole cell lysateab137872 immunoprecipitating LEF1 in Jurkat whole cell lysate. For western blotting, primary antibody used was ab137872 at 1:1000 dilution. ab131366 VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.
Blocking and diluting buffer: 5% NFDM/TBST
Exposure time: 3 minutes
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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Intracellular flow cytometric analysis of Jurkat cell line (human T cell leukemia T lymphocyte) fixed with 4% paraformaldehyde and permeabilized with 90% methanol labeling LEF1 with ab137872 at 1/600 dilution (red). This is compared with a Rabbit monoclonal IgG (ab172730) - Isotype control (black) and a unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti-rabbit IgG (Alexa Fluor® 488) was used as the secondary antibody.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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Immunohistochemical staining of paraffin embedded rat spleen with purified ab137872 at a working dilution of 1/500. The secondary antibody used is ab97051, an HRP-conjugated goat anti-rabbit IgG (H+L), at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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Immunohistochemical staining of paraffin-embedded human thymus with purified ab137872 at a working dilution of 1/500. The secondary antibody used is ab97051, an HRP-conjugated goat anti-rabbit IgG (H+L), at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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Immunohistochemical staining of paraffin embedded human tonsil with purified ab137872 at a working dilution of 1/500. The secondary antibody used is ab97051, an HRP-conjugated goat anti-rabbit IgG (H+L), at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
ab137872 staining LEF1 in paraffin embedded human thymona tissue by Immunohistochemistry. Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins. Samples were incubated with primary antibody at 1/2000 dilution for 30 mins at room temperature. A ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Hematoxylin was used as a counterstain. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. -
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
ab137872 staining LEF1 in paraffin embedded human melanoma tissue by Immunohistochemistry. Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins. Samples were incubated with primary antibody at 1/2000 dilution for 30 mins at room temperature. A ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Hematoxylin was used as a counterstain. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. -
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab137872).
Tissue Microarrays stained for Anti-LEF1 antibody [EPR2029Y] using ab137872 in immunohistochemical analysis. This table provides a detailed overview of positive (tick mark) and negaive (cross mark) staining per sample type tested. The sections were incubated with ab137872 for 30 mins at room temperature used at 1:2000 dilution (1.05 μg/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB secondary antibody (ab209101). Counterstain was Hematoxylin.
Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument.
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This IHC data was generated using the same anti-LEF1 antibody clone, EPR2029Y, in a different buffer formulation (cat# ab137872).
Ab137872 staining LEF1 in paraffin embedded Mouse spleen tissue by Immunohistochemistry. Antigen retrieval was by heat mediation using ab93684 (Tris/EDTA buffer, Ph9). Samples were incubated with primary antibody at 1:2000 dilution. A ready to use Goat Anti-rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counter stain. Nuclear staining on T cells in periarterial lymphatic sheath of mouse spleen (PMID: 21685909).
Protocols
Datasheets and documents
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Datasheet download
Certificate of Compliance
References (3)
ab215999 has been referenced in 3 publications.
- Li J et al. The oncogenic role of Wnt10a in colorectal cancer through activation of canonical Wnt/ß-catenin signaling. Oncol Lett 17:3657-3664 (2019). PubMed: 30881490
- Skokowa J et al. Interactions among HCLS1, HAX1 and LEF-1 proteins are essential for G-CSF-triggered granulopoiesis. Nat Med 18:1550-9 (2012). PubMed: 23001182
- Tandon B et al. Nuclear overexpression of lymphoid-enhancer-binding factor 1 identifies chronic lymphocytic leukemia/small lymphocytic lymphoma in small B-cell lymphomas. Mod Pathol 24:1433-43 (2011). IHC ; Human . PubMed: 21685909