Revision 1

#9383Store at -20C

1 Kit

(4 x 20 microliters)

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Mol. Wt Isotype/Source
TCF1/TCF7 (C63D9) Rabbit mAb 2203 20 µl 48, 50 kDa Rabbit IgG
TCF3/TCF7L1 (D15G11) Rabbit mAb 2883 20 µl 78 kDa Rabbit 
TCF4/TCF7L2 (C48H11) Rabbit mAb 2569 20 µl 58, 79 kDa Rabbit IgG
LEF1 (C12A5) Rabbit mAb 2230 20 µl 25-58 kDa Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl Goat 

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The TCF/LEF Family Antibody Sampler Kit provides an economical means of detecting total protein from the TCF/LEF1 family members. The kit contains enough primary and secondary antibodies to perform two Western blots with each antibody.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Background

LEF1 and TCF are members of the high mobility group (HMG) DNA binding protein family of transcription factors which consists of the following: Lymphoid Enhancer Factor 1 (LEF1), T Cell Factor 1 (TCF1, also known as TCF7), TCF3 (also known as TCF7L1) and TCF4 (also known as TCF7L2) (1). LEF1 and TCF1/TCF7 were originally identified as important factors regulating early lymphoid development (2) that act downstream in Wnt signaling. LEF1 and TCF bind to Wnt response elements to provide docking sites for β-catenin, which translocates to the nucleus to promote the transcription of target genes upon activation of Wnt signaling (3). LEF1 and TCF proteins are dynamically expressed during development and aberrant activation of the Wnt signaling pathway is involved in many types of cancers including colon cancer (4,5).

  1. Waterman, M.L. (2004) Cancer Metastasis Rev 23, 41-52.
  2. Schilham, M.W. and Clevers, H. (1998) Semin Immunol 10, 127-32.
  3. Brantjes, H. et al. (2002) Biol Chem 383, 255-61.
  4. Reya, T. and Clevers, H. (2005) Nature 434, 843-50.
  5. Logan, C.Y. and Nusse, R. (2004) Annu Rev Cell Dev Biol 20, 781-810.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

    Limited Uses

    Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

    Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

    Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
    For Research Use Only. Not for Use in Diagnostic Procedures.
    Revision 1
    #9383

    TCF/LEF Family Antibody Sampler Kit

    TCF/LEF Family Antibody Sampler Kit: Image 1 Expand Image
    Confocal immunofluorescent analysis of fixed frozen lymph node from wild-type (left) Tcf7GFP flox (right; Jax Strain 030909) mice using TCF1/TCF7 (C63D9) Rabbit mAb #2203 (red) and CD4 (RM4-5) Rat mAb (redFluor™ 710 Conjugate) #75508 (blue). EGFP insertion around Tcf7 exon 2 interferes with expression of long isoforms, but not short isoforms.
    TCF/LEF Family Antibody Sampler Kit: Image 2 Expand Image

    Flow cytometric analysis of human peripheral blood mononuclear cells using LEF1 (C12A5) Rabbit mAb (right) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (left), co-stained with CD3 (UCHT1) Mouse mAb (APC Conjugate) #19881. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.

    TCF/LEF Family Antibody Sampler Kit: Image 3 Expand Image
    Simple Western™ analysis of lysates (0.1 mg/mL) from HepG2 cells using TCF4/TCF7L2 (C48H11) Rabbit mAb #2569. The virtual lane view (left) shows the target bands (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ ​​​​​​​Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.

    TCF/LEF Family Antibody Sampler Kit: Image 4 Expand Image
    Western blot analysis of total cell lysates from HT29, Colo201, Jurkat and mouse thymocytes using TCF1/TCF7 (C63D9) Rabbit mAb.
    TCF/LEF Family Antibody Sampler Kit: Image 5 Expand Image
    Western blot analysis of total cell lysates from HCT15, DLD1 and mouse thymocytes using LEF1 (C12A5) Rabbit mAb.
    TCF/LEF Family Antibody Sampler Kit: Image 6 Expand Image
    Western blot analysis of extracts from various cell types using TCF4/TCF7L2 (C48H11) Rabbit mAb.
    TCF/LEF Family Antibody Sampler Kit: Image 7 Expand Image
    CUT&Tag was performed with HCT 116 cells and TCF4/TCF7L2 (C48H11) Rabbit mAb, using CUT&Tag Assay Kit #77552. DNA library was prepared using CUT&Tag Dual Index Primers and PCR Master Mix for Illumina Systems #47415. The figure shows binding across MYC, a known target gene of TCF4 (see our ChIP-qPCR figure).
    TCF/LEF Family Antibody Sampler Kit: Image 8 Expand Image
    Western blot analysis of total lysates from various cell lines using TCF3/TCF7L1 (D15G11) Rabbit mAb.
    TCF/LEF Family Antibody Sampler Kit: Image 9 Expand Image
    After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
    TCF/LEF Family Antibody Sampler Kit: Image 10 Expand Image
    Immunohistochemical analysis of paraffin-embedded human Non-Hodgkin lymphoma using TCF1/TCF7 (C63D9) Rabbit mAb.
    TCF/LEF Family Antibody Sampler Kit: Image 11 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HCT116 cells and either TCF4/TCF7L2 (C48H11) Rabbit mAb or Non-phospho (Active) β-Catenin (Ser33/37/Thr41) (D13A1) Rabbit mAb #8814, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. TCF4/TCF7L2 and β-Catenin are known to associate with each other on chromatin. The figure shows binding of both TCF4/TCF7L2 and β-Catenin across CAMK2D, a known target gene of both TCF4/TCF7L2 and β-Catenin (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product datasheet.
    TCF/LEF Family Antibody Sampler Kit: Image 12 Expand Image
    CUT&Tag was performed with HCT116 cells and TCF4/TCF7L2 (C48H11) Rabbit mAb, using CUT&Tag Assay Kit #77552. DNA library was prepared using CUT&Tag Dual Index Primers and PCR Master Mix for Illumina Systems #47415. The figures show binding across chromosome 8 (upper), including MYC (lower), a known target gene of TCF4 (see our ChIP-qPCR figure).
    TCF/LEF Family Antibody Sampler Kit: Image 13 Expand Image
    Immunohistochemical analysis of paraffin-embedded human lung carcinoma using TCF1/TCF7 (C63D9) Rabbit mAb in the presence of control peptide (left) or TCF1/TCF7 blocking peptide #1007 (right).
    TCF/LEF Family Antibody Sampler Kit: Image 14 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HCT116 cells and either TCF4/TCF7L2 (C48H11) Rabbit mAb or Non-phospho (Active) β-Catenin (Ser33/37/Thr41) (D13A1) Rabbit mAb #8814, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. TCF4/TCF7L2 and β-Catenin are known to associate with each other on chromatin. The figure shows binding of both TCF4/TCF7L2 and β-Catenin across chromosome 4 (upper), including CAMK2D (lower), a known target gene of both TCF4/TCF7L2 and β-Catenin (see additional figure containing ChIP-qPCR data).
    TCF/LEF Family Antibody Sampler Kit: Image 15 Expand Image
    Immunohistochemical analysis of paraffin-embedded human tonsil using TCF1/TCF7 (C63D9) Rabbit mAb.
    TCF/LEF Family Antibody Sampler Kit: Image 16 Expand Image
    Confocal immunofluorescent analysis of HCT-15 cells using LEF1 (C12A5) Rabbit mAb (green). Actin filaments have been labeled with DyLight™ 554 Phalloidin #13054 (red).
    TCF/LEF Family Antibody Sampler Kit: Image 17 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HCT 116 cells and either TCF4/TCF7L2 (C48H11) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human CaMK2D Intron 3 Primers #5111, human c-Myc promoter primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    TCF/LEF Family Antibody Sampler Kit: Image 18 Expand Image
    Confocal immunofluorescent analysis of DLD-1 cells using TCF1/TCF7 (C63D9) Rabbit mAb (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin (red).
    TCF/LEF Family Antibody Sampler Kit: Image 19 Expand Image

    Flow cytometric analysis of mouse splenocytes using LEF1 (C12A5) Rabbit mAb (right) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (left), co-stained with CD3 (17A2) Rat mAb (APC Conjugate) #24265. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.

    TCF/LEF Family Antibody Sampler Kit: Image 20 Expand Image
    CUT&RUN was performed with HCT 116 cells and TCF4/TCF7L2 (C48H11) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across MYC, a known target gene of TCF4 (see additional figure containing CUT&RUN-qPCR data).
    TCF/LEF Family Antibody Sampler Kit: Image 21 Expand Image
    Flow cytometric analysis of A20 cells (blue) and EL-4 cells (green) using TCF1/TCF7 (C63D9) Rabbit mAb (solid lines) or a concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
    TCF/LEF Family Antibody Sampler Kit: Image 22 Expand Image
    CUT&RUN was performed with HCT 116 cells and TCF4/TCF7L2 (C48H11) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 8 (upper), including MYC (lower), a known target gene of TCF4 (see additional figure containing CUT&RUN-qPCR data).
    TCF/LEF Family Antibody Sampler Kit: Image 23 Expand Image
    CUT&RUN was performed with HCT 116 cells and either TCF4/TCF7L2 (C48H11) Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using human PRKACB intron 1 primers, human MYC exon 1 primers, and human ITM2A upstream primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    TCF/LEF Family Antibody Sampler Kit: Image 24 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from mouse thymocytes and either TCF1/TCF7 (C63D9) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. The enriched DNA was quantified by real-time PCR using SimpleChIP® Mouse LEF1 Upstream Primers #80993 and SimpleChIP® Mouse MYT-1 Promoter Primers #8985. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
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    Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
    For Research Use Only. Not for Use in Diagnostic Procedures.