Revision 12
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.
Applications:

WB, W-S, IP, IHC-Bond, IHC-P, IF-F, IF-IC, FC-FP, ChIP, ChIP-seq, C&R

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

92

Source/Isotype:

Rabbit IgG

UniProt ID:

#P35222

Entrez-Gene Id:

1499

Product Information

Product Usage Information

For optimal ChIP and ChIP-seq results, use 20 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

The CUT&RUN dilution was determined using CUT&RUN Assay Kit #86652.
Application Dilution
Western Blotting 1:1000
Simple Western™ 1:10 - 1:50
Immunoprecipitation 1:50
IHC Leica Bond 1:50 - 1:200
Immunohistochemistry (Paraffin) 1:50 - 1:200
Immunofluorescence (Frozen) 1:50 - 1:100
Immunofluorescence (Immunocytochemistry) 1:100 - 1:400
Flow Cytometry (Fixed/Permeabilized) 1:50 - 1:100
Chromatin IP 1:25
Chromatin IP-seq 1:25
CUT&RUN 1:25

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.

For a carrier free (BSA and azide free) version of this product see product #84441.

Specificity / Sensitivity

β-Catenin (D10A8) XP® Rabbit mAb recognizes endogenous levels of total β-catenin protein.

Species Reactivity:

Human, Mouse, Rat, Monkey

Species predicted to react based on 100% sequence homology

Zebrafish, Bovine, Pig, Horse, Guinea Pig

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro714 of human β-catenin protein.

Background

β-catenin is a key downstream effector in the Wnt signaling pathway (1). It is implicated in two major biological processes in vertebrates: early embryonic development (2) and tumorigenesis (3). CK1 phosphorylates β-catenin at Ser45. This phosphorylation event primes β-catenin for subsequent phosphorylation by GSK-3β (4-6). GSK-3β destabilizes β-catenin by phosphorylating it at Ser33, Ser37, and Thr41 (7). Mutations at these sites result in the stabilization of β-catenin protein levels and have been found in many tumor cell lines (8).

  1. Cadigan, K.M. and Nusse, R. (1997) Genes Dev 11, 3286-3305.
  2. Wodarz, A. and Nusse, R. (1998) Annu Rev Cell Dev Biol 14, 59-88.
  3. Polakis, P. (1999) Curr Opin Genet Dev 9, 15-21.
  4. Amit, S. et al. (2002) Genes Dev 16, 1066-76.
  5. Liu, C. et al. (2002) Cell 108, 837-47.
  6. Yanagawa, S. et al. (2002) EMBO J 21, 1733-42.
  7. Yost, C. et al. (1996) Genes Dev 10, 1443-54.
  8. Morin, P.J. et al. (1997) Science 275, 1787-90.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting W-S: Simple Western™ IP: Immunoprecipitation IHC-Bond: IHC Leica Bond IHC-P: Immunohistochemistry (Paraffin) IF-F: Immunofluorescence (Frozen) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq C&R: CUT&RUN

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SignalStain is a registered trademark of Cell Signaling Technology, Inc.
SimpleChIP is a registered trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

使用に関する制限

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Revision 12
#8480

β-Catenin (D10A8) XP® Rabbit mAb

Western Blotting Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from control HeLa cells (lane 1) or HeLa cells with an apparent in-frame truncation mutation in the gene encoding β-Catenin (lane 2) using β-Catenin (D10A8) XP® Rabbit mAb, #8480 (upper) or β-actin (D6A8) Rabbit mAb #8457 (lower). The change in β-Catenin molecular weight in the mutated HeLa cells is consistent with an in-frame deletion.
Western Blotting Image 2: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from various cell lines using β-Catenin (D10A8) XP® Rabbit mAb.
Western Blotting Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Simple Western™ analysis of lysates (0.1 mg/mL) from HeLa cells using β-Catenin (D10A8) XP® Rabbit mAb #8480. The virtual lane view (left) shows the target band (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.
Immunoprecipitation Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunoprecipitation of β-Catenin from HeLa cell extracts. Lane 1 is 10% input, lane 2 is precipitated with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is β-Catenin (D10A8) XP® Rabbit mAb, #8480. Western blot was performed using β-Catenin (15B8) Mouse mAb, #37477.
Immunohistochemistry Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human prostate adenocarcinoma using ß-Catenin (D10A8) XP® Rabbit mAb performed on the Leica® BOND Rx.
Immunohistochemistry Image 2: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human colon adenocarcinoma using ß-Catenin (D10A8) XP® Rabbit mAb performed on the Leica® BOND Rx.
Immunohistochemistry Image 3: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human serous adenocarcinoma of the ovary using ß-Catenin (D10A8) XP® Rabbit mAb performed on the Leica® BOND Rx.
Immunohistochemistry Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human colon adenocarcinoma using ß-Catenin (D10A8) XP® Rabbit mAb.

Immunohistochemistry Image 2: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human lung carcinoma using β-Catenin (D10A8) XP® Rabbit mAb.
Immunohistochemistry Image 3: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human colon adenocarcinoma using ß-Catenin (D10A8) XP® Rabbit mAb.
Immunohistochemistry Image 4: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human colon carcinoma using β-Catenin (D10A8) XP® Rabbit mAb.
Immunohistochemistry Image 5: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human breast carcinoma using β-Catenin (D10A8) XP® Rabbit mAb.
Immunohistochemistry Image 6: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded mouse colon using β-Catenin (D10A8) XP® Rabbit mAb in the presence of control peptide (left) or antigen-specific peptide (right).
Immunohistochemistry Image 7: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded cell pellets, HeLa (left) or NCI-H28 (right), using β-Catenin (D10A8) XP® Rabbit mAb.
Immunofluorescence Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of mouse colon using β-Catenin (D10A8) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Immunofluorescence Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of HeLa (left) and NCI-H28 (right) cells using β-Catenin (D10A8) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Flow Cytometry Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Flow cytometric analysis of NCI-H28 cells (green) and HeLa cells (blue) using β-Catenin (D10A8) XP® Rabbit mAb (solid lines) or concentration-matched Rabbit Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as secondary antibody.
Chromatin Immunoprecipitation Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from HCT116 cells and either β-Catenin (D10A8) XP® Rabbit mAb or Non-phospho (Active) β-Catenin (Ser33/37/Thr41) (D13A1) Rabbit mAb #8814, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across AXIN2, a known target gene of β-Catenin (see additional figure containing ChIP-qPCR data).
Chromatin Immunoprecipitation Image 2: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from HCT116 cells and either β-Catenin (D10A8) XP® Rabbit mAb or Non-phospho (Active) β-Catenin (Ser33/37/Thr41) (D13A1) Rabbit mAb #8814, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across chromosome 17 (upper), including AXIN2 (lower), a known target gene of β-Catenin (see additional figure containing ChIP-qPCR data).
Chromatin Immunoprecipitation Image 3: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from HCT 116 cells and either β-Catenin (D10A8) XP® 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 Axin2 Intron 1 Primers #8973, 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.
CUT and RUN Image 1: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
CUT&RUN was performed with HCT 116 cells and β-Catenin (D10A8) XP® Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA library was prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across Axin2, a known target gene of β-Catenin (see additional figure containing CUT&RUN-qPCR data).
CUT and RUN Image 2: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
CUT&RUN was performed with HCT 116 cells and β-Catenin (D10A8) XP® 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 17 (upper), including Axin2 (lower), a known target gene of β-Catenin (see additional figure containing CUT&RUN-qPCR data).
CUT and RUN Image 3: β-Catenin (D10A8) XP® Rabbit mAb Expand Image
CUT&RUN was performed with HCT 116 cells and either β-Catenin (D10A8) XP® 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 SimpleChIP® Human Axin2 Intron 1 Primers #8973, SimpleChIP® Human CaMK2D Intron 3 Primers #5111 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.