33-4000

antibody from Invitrogen Antibodies

Targeting: CDH1 CD324, UVO, uvomorulin

Provider product page for 33-4000

Western blot

Immunoprecipitation

Immunohistochemistry

Other assay

Antibody data

Antibody Data
Antigen structure
References [213]
Comments [0]
Validations
Western blot [2]
Other assay [9]

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Product number: 33-4000 - Provider product page
Provider: Invitrogen Antibodies
Product name: E-cadherin Monoclonal Antibody (4A2C7)
Antibody type: Monoclonal
Antigen: Recombinant full-length protein
Description: This antibody is specific for E-cadherin but does show some minor cross-reactivity with P-cadherin. Formalin-fixed, paraffin embedded tissue sections required a Heat Induced Epitope Retrieval (HIER) step prior to staining with this antibody in an IHC application.
Reactivity: Human
Host: Mouse
Isotype: IgG
Antibody clone number: 4A2C7
Vial size: 100 µg
Concentration: 0.5 mg/mL
Storage: -20°C

CDH1 protein structure - 33-4000 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of E-Cadherin was performed by loading 20 µg of A431 (lane1) and Rat Thyroid lysate (lane2) lysates using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (LC5800). Proteins were transferred to a PVDF membrane and blocked with 5 % skim milk for 1 hour at room temperature. E-Cadherin was detected at ~110 kDa using E-Cadherin Mouse Monoclonal Antibody (Product # 33-4000) at 1-3 µg/mL in 2.5 % skim milk at 4°C overnight on a rocking platform. Goat Anti-Mouse IgG - HRP Secondary Antibody (Product # 62-6520) at 1:4000 dilution was used and chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-E-cadherin Monoclonal Antibody (4A2C7) (Product # 33-4000) and a 110kDa band corresponding to E-cadherin was observed across in all tested cell lines and tissues, except MDA-MB-231, SW480 and BJ. Whole cell extracts (30 µg lysate) of MCF7 (Lane 1), MDA-MB-231 (Lane 2), SW480 (Lane 3), Caco-2 (Lane 4), BJ (Lane 5), Mouse Stomach (Lane 6), Rat Stomach (Lane 7) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody ( 1 µg/mL) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177,1:4000) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).The band at ~80kDa in Mouse stomach and Rat stomach corresponds to proteolytic cleavage products of E-cadherin.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 Effects of suppression of cofilin activation or overexpresssion of TJ proteins on the capsaicin-induced cofilin dephosphorylation and decrease in occludin expression. (A-C) Flag-HA tagged cofilin and LIMK, and EGFP-occludin and claudin-1 stable transfectants were established. Monolayers prepared from the transfectants and non-transfected MDCK cells were exposed to 300 uM capsaicin for the time indicated. After lysis, protein expression was analyzed by immunoblotting. Each experiment was performed with at least two different clones and repeated at least twice. (D) MDCK monolayers were pretreated with vehicle or 50 uM TFP for 30 min, and then exposed to 100 uM capsaicin for the time indicated. The levels of occludin and phospho-cofilin were analyzed by western blotting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 7 Cofilin activation, decrease in occludin expression and actin alteration in the recovery phase of capsaicin treatment. (A) Western blot detection of phospho-cofilin and occludin in total extracts exposed to 300 uM capsaicin for the times indicated. (B) MDCK monolayers were exposed to 300 uM capsaicin for the time indicated, and then fixed and stained with rhodamine-phalloidin to detect F-actin. Images from each z-section were deconvoluted and overlayed. Bar: 10 um.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3 Effects of capsaicin on the distribution of TJ proteins and F-actin. (A, B) Monolayers were exposed to vehicle (A) or 300 uM capsaicin (B) for 45 min and were labeled with each TJ antibody (green), rhodamine-phalloidin (red) and Hoechst (blue). Images were collected as a Z-series, and then deconvoluted and overlayed to display a single composite projection. Top : XY sections of merged images. Bottom : XZ sections of merged images. Scale bar: 10 um. The corresponding TJ antibodies are listed above the images. (C) 3D projection images in a 45deg-angle of claudin-1 and tricellulin staining, plus F-actin. Scale bar: 10 um. (D) MDCK monolayers exposed to vehicle (top) or capsaicin (bottom) as above were stained with E-cadherin. The colors corresponding to each antibody are listed above the images. Scale bar: 2.5 um. (E) Schematic explanation of the protein distributions.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 Claudin-15 and ion transport are modulated in Hnf4alpha colon mutants. (A) qRT-PCR of claudin-4, -8 and -15 at various time points in mutants (Open bars) relative to controls (Black bars). Expression levels are shown as mean values (+- SEM) relative to controls at each time point (3-14 mice per group) and were normalized with the Tbp housekeeping gene. (B) Immunoblotting confirmed modulated protein levels of claudin-4, -8 and -15 at 12 months in mutants (n = 3, open bars) and relative controls (n = 3, black bars) normalized to stable expression of E-Cadherin (Cdh1). (C) Quantification of band intensities of each claudin (means +- SEM; n = 3 or 4 for each group) relative to Cdh1. (D) Indirect immunofluorescence (FITC) of claudin-4, -8 and -15 in 3 month-old animals in proximal colon (representative of 4 pairs). Nuclei were counterstained with DAPI (blue) and merged with FITC (green). Scale bars, 50 um. (E) Claudin-15 expression is lost in the upper half of the colon epithelium in mutants (Open bar) relative to controls (Black bar). (F) Colonic permeability to 51 Cr-EDTA measured in colon tissues from 2 month-old mice (5 mice per group). (G) Baseline short circuit current (Isc) measured in colon tissues from 2 month-old mice (5 mice per group). Represented as mean values +- SEM, * P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 4 Capsaicin decreases occludin protein content and the protein-protein interactions involving occludin in TJ. (A,B) MDCK monolayers were exposed to ethanol control or 300 uM capsaicin for 45 min. (A) The cellular localization of the TJ proteins occludin and claudin-1 was examined by immunofluorescence. Scale bar: 10 um. (B) Western blot detection of occludin, tricellulin, Zo-1, claudin-1, E-cadherin and actin in the cytosol, membrane fractions and total cell extracts. The densitometic analysis of total proteins from three independent experiments performed with NIH ImageJ software. **represents p

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 Transient conditional depletion of Foxp3 + Tregs activates the CP for leukocyte trafficking and mitigates AD pathology. ( a ) AD-Tg/Foxp3-DTR + and non-DTR-expressing AD-Tg littermates (AD-Tg/Foxp3-DTR - ; AD-Tg control) were treated with DTx for 4 consecutive days. mRNA levels of icam1 , cxcl10 and ccl2 in the CP of 6-month-old AD-Tg/Foxp3-DTR + mice, 1 day after last injection ( n =6-8 per group; Student's t -test). ( b ) Representative images of the CP, immunostained for E-Cadherin (green), ICAM-1 (red) and Hoechst, in 6-month-old AD-Tg/Foxp3-DTR + mice, 1 day after last DTx injection (Scale bar, 25 mum). ( c ) mRNA levels of icam1 and vcam1 , in cultured CP cells, 24 h after addition of ex vivo differentiated Tregs, IFN-gamma or their combination, relative to untreated (UT) cells ( n =3-4 per group; one-way ANOVA followed by Newman-Keuls post hoc test; NS, not significant). ( d - f ) Flow cytometry analysis of the brain of 6-month-old AD-Tg/Foxp3-DTR + mice, 3 weeks following last DTx injection, showing increased numbers of CD11b high /CD45 high mo-MPhi and CD4 + T cells ( d ), and increased CD4 + Foxp3 + ( e , f ) Treg frequencies ( n =4-7 per group; Student's t -test). ( g ) mRNA levels of foxp3 and il10 in the brain of 6-month-old AD-Tg/Foxp3-DTR + mice, 3 weeks after last DTx injection ( n =6-8 per group; Student's t -test). ( h , i ) Representative images of Abeta (green) and CD3 (red) ( h ), or Foxp3 and IL-10 (in red) immunostaining ( i ) in