After 72 hours of non–T- or PAICS-siRNA transfection, the RT112 cells were seeded in chamber slides (Lab-Tek II CC2, Nunc, Rochester, NY) and incubated for a day to immunostain with rabbit monoclonal E-cadherin antibody (IF, 1:200; catalog #3195, Cell Signaling Technology, Danvers, MA). The slides were washed with phosphate-buffered saline (PBS) and were fixed using ice-cold methanol. Following three times PBS with 0.05% Tween 20 (PBS-T) wash, the slides were blocked for 2 hours using 5% normal horse serum in PBS-T. A rabbit anti-E-cadherin antibody was added to the slides at 1:200 dilution and incubated for 1 hour at room temperature. Following three times PBS-T wash, the slides were incubated with Alexa 555–conjugated goat, anti-rabbit antibody (Invitrogen) for 1 hour in the dark at room temperature. After three times PBS wash, the slides were mounted using ProLong Gold Antifade Mountant with DAPI (catalog #P36931, Invitrogen, ThermoFisher Scientific, Carlsbad, CA). Confocal images were taken with a 60× lens Nikon A1 High Speed Laser Confocal Spectral Imaging microscope (Nikon Instruments Inc., Melville, NY) from UAB high-resolution imaging facility.
E cadherin rabbit monoclonal antibody
Manufactured by Cell Signaling Technology
Sourced in United States
The E-cadherin rabbit monoclonal antibody is a laboratory tool used for the detection and analysis of the E-cadherin protein in biological samples. E-cadherin is a cell-cell adhesion molecule that plays a crucial role in the maintenance of cell-cell junctions and epithelial tissue integrity.
Automatically generated - may contain errors
Lab products found in correlation
Prolong gold antifade mountant with dapi, by Thermo Fisher Scientific (1 mentions)
Lab tek 2 cc2, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Lsm780 uv, by Zeiss (1 mentions)
Zo 1 rabbit polyclonal antibody, by Abcam (1 mentions)
Ecl substrate, by Merck Group (1 mentions)
Mini protean tetra cell system, by Bio-Rad (1 mentions)
Immobilon, by Merck Group (1 mentions)
β actin specific antibody, by Merck Group (1 mentions)
Mmp2 rabbit monoclonal antibody, by Cell Signaling Technology (1 mentions)
Mmp 9 rabbit monoclonal antibody, by Cell Signaling Technology (1 mentions)
4 6 diamidino 2 phenylindole dapi, by Abcam (1 mentions)
Rabbit anti adiponectin polyclonal antibody, by Abcam (1 mentions)
6 protocols using e cadherin rabbit monoclonal antibody
1
Immunostaining of E-cadherin in RT112 Cells
2
Investigating Hypoxia-Induced Epithelial-Mesenchymal Transition
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p-Cresyl sulfate (pCS) was obtained from Sigma (Sigma-Aldrich, St. Louis, MO, USA). The Millicell cell culture chamber was obtained from Millipore (Millipore, Billerica, MA, USA). Rabbit polyclonal HIF1α antibody (1:1000) was obtained from Genetex (Alton Parkway, CA, USA). Rabbit monoclonal HIF2α antibody (1:1000), rabbit polyclonal VHL antibody (1:1000), rabbit monoclonal E-cadherin antibody (1:1000), rabbit monoclonal vimentin antibody (1:1000), and rabbit polyclonal α-tubulin antibody (1:5000) were from Cell Signaling Technology (Danvers, MA, USA). Rabbit polyclonal fibronectin antibody (1:2000) and mouse monoclonal twist antibody (1:500) were from Abcam (Cambridge, UK). Secondary antibodies (anti-mouse IgG horseradish peroxidase-linked and anti-rabbit IgG horseradish peroxidase-linked) were form Cell Signaling Technology. Primers were synthesize by Integrated DNA Technologies (Coralville, IA, USA).
3
Immunofluorescence Imaging of EMT Markers in HPV-16 Cells
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Immunofluorescence (IF) staining was used to detect EMT markers expression and localization in CSC-exposed HPV-16 Ect1/E6E7 cells. HPV-16 Ect1/E6E7 cells treated with CSC (3R4F (1 × 10−3, and 10 μg/mL)) for 72 h were fixed and incubated with primary antibodies to E-Cadherin rabbit monoclonal antibody (1:200, Cell Signaling, #31950) and Vimentin mouse monoclonal antibody (1:500, Santa Cruz, sc-6260) at 4 °C overnight, followed by incubation with Alexa Fluor 488 goat anti-Mouse (red fluorescence) secondary antibody (1:5000, Molecular Probes®, Eugene, OR, USA, A11001) and Alexa Fluor 555 goat anti-Rabbit (green fluorescence) secondary antibody (1:5000, Molecular Probes®, A21428) at room temperature for 1 h. After counterstaining with DAPI (CAT# 1306, Molecular Probes) for 30 min, slides were examined under a Zeiss LSM780-UV and LSM880-UV meta confocal microscope (Carl Zeiss Inc.) using a Plan-Apochromat 40′/1.3 Oil DIC objective. Fluorescence intensities and percent of positive cells were measured by ImageJ/Fiji (NIHV 2.9.0/1.54b, National Institutes of Health, USA).
4
Immunohistochemical Profiling of Tissue Markers
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Formalin-fixed tissue was embedded in paraffin and stained with F4/80 rat monoclonal antibody (Novus, NB600-404), CD11b rabbit polyclonal antibody (Novus Cat# NB110-89474), Ly-6G/Ly-6C rat monoclonal antibody (Novus Cat# NBP2-00441), E-cadherin rabbit monoclonal antibody (Cell Signaling Technology Cat# 3195), ZO-1 rabbit polyclonal antibody (Abcam Cat# ab59720), Occludin rabbit polyclonal antibody (Novus Cat# NBP1-87402) and Claudin-1 rabbit polyclonal antibody (Abcam Cat# ab15098).
5
Western Blot Analysis of EMT Markers
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Cultured or transfected cells were lysed in RIPA buffer with 1% PMSF. Western blot was performed on 10% SDS-PAGE by using Mini-PROTEAN® Tetra Cell Systems (Bio-Rad). Proteins were transferred onto polyvinylidene difluoride membranes (Immobilon, Millipore). Membranes were incubated overnight with ITGB1 rabbit monoclonal antibody (Cell Signaling), E-cadherin rabbit monoclonal antibody (Cell Signaling), N-cadherin rabbit monoclonal antibody (Abcam), Vimentin rabbit monoclonal antibody (Abcam), MMP9 rabbit monoclonal antibody (Cell Signaling), or MMP2 rabbit monoclonal antibody (Cell Signaling) at 1 : 1000 dilution or β-actin-specific antibody (Sigma-Aldrich) at 1 : 5000 dilution at 4°C. Signals were visualised using ECL substrates (Millipore, MA, USA).
6
Bicolor Immunofluorescence for Placental Proteins
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Bicolor immunofluorescence was used to identify and localize proteins in sections of WT and NOD mouse placenta tissues. Placentas were fixed in 4% paraformaldehyde, embedded in optimal cutting temperature compound, and sectioned at 5 μm. After deparaffinization, rehydration, and unmasking, slides were blocked with 5% FBS for 1 hr at room temperature. After the blocking buffer was aspirated, sections were incubated with a primary antibody [rabbit anti‐adiponectin polyclonal antibody (1:50 dilution; Abcam) or mouse anticytokeratin 7 monoclonal antibody (1:100 dilution; Abcam)] overnight at 4°C. Cytokeratin 7 was used as a trophoblast marker as previously described.24 After being washed three times with PBS, specimens were incubated with secondary fluorescent antibodies for 1 hr at room temperature in the dark. Sections were finally mounted with fluoroshield mounting medium with 4′,6‐diamidino‐2‐phenylindole (DAPI; Abcam) and then observed using a fluorescence microscope. For the BeWo cells, identification and localization of proteins were conducted using an immunofluorescence assay as previously reported.25 rabbit anti‐adiponectin polyclonal antibody (1:50 dilution; Abcam) and E‐Cadherin rabbit monoclonal antibody (1:200 dilution; Cell Signaling Technology) were used as primary antibodies.
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