50-2567-42

antibody from Invitrogen Antibodies

Targeting: CTNNB1 armadillo, beta-catenin, CTNNB

Provider product page for 50-2567-42

Flow cytometry

Other assay

Antibody data

Antibody Data
Antigen structure
References [4]
Comments [0]
Validations
Flow cytometry [1]
Other assay [7]

Submit

Product number: 50-2567-42 - Provider product page
Provider: Invitrogen Antibodies
Product name: beta Catenin Monoclonal Antibody (15B8), eFluor™ 660, eBioscience™
Antibody type: Monoclonal
Antigen: Other
Description: Description: The 15B8 monoclonal antibody reacts with human and mouse beta-catenin, one member of a family of catenins, which are intracellular proteins that interact with cadherins to mediate cellular adhesion. More specifically, beta-catenin binds to the cytoplasmic tail of E-cadherin. In addition, this molecule is a component of the canonical Wnt signaling pathway. In the absence of Wnt binding its receptor, beta-catenin is phosphorylated and resides in the cytoplasm where it is eventually targeted for degradation by ubiquitination. Upon Wnt binding, beta-catenin becomes dephosphorylated, translocates to the nucleus, and modulates gene expression in partnership with the transcription factors T cell factor (TCF) and lymphocyte enhancer binding factor (LEF). Expression of beta-catenin is found in a wide variety of non-immune and immune tissues, including thymocytes and T and B lymphocytes. The Wnt and beta-catenin signaling pathway has been demonstrated to play a crucial role in the development of T, B, and hematopoietic stem cells. Applications Reported: This 15B8 antibody has been reported for use in intracellular staining followed by flow cytometric analysis. Applications Tested: This 15B8 antibody has been tested by intracellular staining and flow cytometric analysis of the Jurkat cell line using the Foxp3/Transcription Factor Staining Buffer Set (Product # 00-5523-00). This can be used at less than or equal to 0.25 µg per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest. eFluor® 660 is a replacement for Alexa Fluor® 647. eFluor® 660 emits at 659 nm and is excited with the red laser (633 nm). Please make sure that your instrument is capable of detecting this fluorochome. Excitation: 633-647 nm; Emission: 668 nm; Laser: Red Laser. Filtration: 0.2 µm post-manufacturing filtered.
Reactivity: Human, Mouse
Host: Mouse
Isotype: IgG
Antibody clone number: 15B8
Vial size: 100 Tests
Concentration: 5 µL/Test
Storage: 4° C, store in dark, DO NOT FREEZE!

CTNNB1 protein structure - 50-2567-42 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 Chemoresistant NSCLC cells exhibit an activated EMT pathway. ( a ) Morphological images of chemoresistant A549_R and parental A549 cells. Scale bar, 100 mu m. ( b ) A549_R and A549 cells were immunostained for Vimentin, beta -catenin and E-cadherin (green). Nuclei were counterstained with DAPI (blue). Scale bar, 25 mu m ( c-e ) mRNA levels of EMT-TFs and CDH2/N-cadherin in A549_R, H358_R and H460_R cells and the corresponding parental cells (A549, H358 and H460) were analyzed by qPCR. Data are shown as mean+-S.D. of three independent experiments ( n =3). ( f ) Different NSCLC cells are at distinct stages along the EMT. mRNA levels of EMT-TFs and Vimentin in PC9, H358, A549 and H460 cells were analyzed by qPCR and results are shown in mean+-S.D. of three independent experiments ( n =3). ( g ) NSCLC cells at distinct EMT stages exhibit differential sensitivity to MTA. PC9, H358, A549 and H460 cells treated with the indicated MTA doses for 5 days were analyzed by XTT assay. The results are shown as mean+-S.D. of triplicate experiments. * P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 1 Wnt signaling alters human memory CD8 + T cell proliferation and differentiation. ( A ) FACS-sorted human naive and memory CD8 + T cells were stained with CTV and stimulated with CD3/CD28 in DMSO (black), TWS119 (blue), and SKL2001 (red) for 7 days. Numbers represent the percentages of divided cells in each treatment. ( B ) Average proliferative response for each treatment ( n = 5). Dunn's test for multiple comparisons. ( C ) Memory CD8 + T cells in different treatments for 7 days were stained with phenotypic markers. Numbers represent the percentages of cells in each quadrant. ( D ) Quantitative RT-PCR analysis of Tcf7, Lef1, and Fzd7 in memory CD8 + T cells with or without Wnt agonist treatment on days 1, 3, and 5. Numbers are normalized to marker expression of the DMSO group at each corresponding time point. ( E ) Memory CD8 + T cells were stimulated with CD3/CD28 and treated with or without Wnt agonists for 7 days. beta-Catenin and TCF1 levels were analyzed by flow cytometry. Numbers represent the MFI of each protein. ( F ) Average MFI of beta-catenin and TCF1 in different treatment conditions ( n = 4). Dunn's test for multiple comparisons. # P < 0.05.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2. Junctional proteins and proteins involved in calcium handling. Overlay images including DAPI are shown on the left, followed by corresponding green and red channel images in greyscale. A) Cells were immunostained for beta-catenin (green), connexin-43 (red) and DNA (blue). B) Cells were immunostained for N-cadherin (green), desmoglein- 2 (red) and DNA (blue). C) Cells were immunostained for BIN-1/amphiphysin-2 (green) and EH-myomesin (red). D) Cells were immunostained for SERCA-2a (green), EHmyomesin (red) and DNA (blue). E) Cells were immunostained for ryanodine receptors (green), EH-myomesin (red) and DNA (blue).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 beta-catenin is related to ALDH HIGH activity cell population . (A,B) Density plots showing the percentage of ALDH HIGH cells in SiHa cell line grown in monolayer or spheres cultures. The bar graph showed the percentages of ALDH HIGH cells in monolayer (white bar) or spheres cultures (black bar). The enzymatic activity of ALDH was measured by flow cytometry using the ALDEFLUOR kit. The inhibitor DEAB was employed as a background fluorescence control (data not shown). Data was normalized with DEAB (internal control) from 3 independent experiments are showed in bar graph (Means +- S.E.M. ), * p < 0.05. (C) Western Blot of beta-catenin in SiHa cells transfected with siRNA scramble (negative control) or siRNA beta-catenin at 100 nM during 24 h. GAPDH was used as loading control. (D) The units were normalized with siRNA scramble and the densitometry quantification from 3 independent experiments are showed in bar graph (Means +- S.E.M), ** p < 0.01. (E,F) Density plots showing the percentage of ALDH HIGH cells obtained of spheres cultures from SiHa cells transfected with siRNA scramble (left panel) or siRNA beta-catenin (right panel). Transient transfection of SiHa cells using siRNA scramble (control) or siRNA beta-catenin was employed to knockdown beta-catenin. At 24 h post-transfection, SiHa cells were growth in spheres cultures for an additional 72 h. Each percentage of ALDH HIGH cells was normalized with DEAB as internal control. The percentages of ALDH HIGH cells with

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 4 GSK3-beta inhibition upregulates ALDH through beta-catenin-dependent transcriptional activity. (A) Images showing beta-catenin (green) localization in SiHa cells treated with DMSO (vehicle) or 5 muM of BIO obtained by immunofluorescence microscopy. The nuclei were stained with IP (red). Representative images from at least 3 independent experiments are showed. Magnification 40X. (B) Reporter activity of beta-catenin-dependent transcription in SiHa cells exposed to BIO treatment. Transient transfection of SiHa cells using the plasmid control pFOP (black bars) and the reporter plasmid pTOP (white bars) was employed to measure transcriptional activity of beta-catenin. At 24 h post-transfection, the SiHa cells were treated with BIO at the concentrations indicated for an additional 48 h. The values of luciferase were normalized to beta-galactosidase and the data are showed in bar graph from 5 independent experiments, (Means +- S.E.M). * p < 0.05, ** p < 0.01. (C,D) Western Blot of ALDH in SiHa cells growth as monolayer culture and exposed to BIO at the concentrations indicated during 48 h. GAPDH was used as loading control. Densitometry quantification is showed in bar graph (Means +- S.E.M). (E,F) Density plots showing the percentage of ALDH HIGH cells in SiHa cells treated with DMSO (vehicle) or BIO. The enzymatic activity of ALDH was measured by flow cytometry using the ALDEFLUOR kit. DEAB inhibitor was employed to determine the percentage of ALDH HIGH cells for each con

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 The ALDH HIGH subpopulation from spheres model shown elements associated to the activation of beta-catenin. (A) Schematic model of cell sorting to ALDH HIGH and ALDH LOW subpopulations from spheres grown 72 h. (B) Western Blot analysis of beta-catenin, p-GSK3-beta Ser9, and p-AKT Ser473 and total AKT from sorted ALDH subpopulations.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 6 The AKT activity is present in upstream beta-catenin/ALDH axis. (A) Representative image of spheres size upon 0.1-2.0 muM AZD-5363 treatment. (B-E) Western Blot analysis of active beta-catenin, p-GSK3-beta Ser9, and ALDH protein from spheres treated with 0.1-2.0 muM AZD-5363, the bar graphs shown the comparation of control (DMSO) vs. 2.0 muM AZD-5363 treatment from 3 independent experiments (Means +- S.E.M. ), * p < 0.05,*** p < 0.001. (F) Density plots showing the percentage of ALDH HIGH cells in spheres treated with control (DMSO) or 2.0 muM AZD-5363 treatment for 72 h. (G) The bar graph shown the percentage of ALDEFLUOR positive cells in both conditions, DMSO and AZD-5363 treatment, considering 3 independent experiments (Means +- S.E.M. ), * p < 0.05.