The back skins from Dct-Lac-Z CD1 transgenic mice were fixed at 4°C in 4% paraformaldehyde for 1 h, and stained with x-gal (Beyotime, China) for 24 h. After wash, the skin specimens were embedded in paraffin and sliced into 5 μm sections, which were incubated with rabbit anti-β-catenin polyclonal antibody (1:600; Abcam, USA) at 4℃ for overnight, and the secondary antibody (Zhongshan, China) at 37°C for 1h. The samples were developed with DAB (Zhongshan, China).
Rabbit anti β catenin polyclonal antibody
Manufactured by Abcam
Sourced in United States
Rabbit anti-β-catenin polyclonal antibody is a laboratory reagent used for the detection and quantification of β-catenin, a key signaling molecule involved in various cellular processes. This antibody is produced in rabbits and recognizes the β-catenin protein.
Automatically generated - may contain errors
Lab products found in correlation
X gal, by Beyotime (1 mentions)
Fluorescence microscope, by Leica (1 mentions)
Permount mounting medium, by Merck Group (1 mentions)
Goat anti rabbit igg fitc, by Abcam (1 mentions)
Goat anti mouse rabbit igg, by Abcam (1 mentions)
Rabbit anti caspase 3 polyclonal antibody, by Abcam (1 mentions)
Mouse anti β actin polyclonal antibody, by Abcam (1 mentions)
4 protocols using rabbit anti β catenin polyclonal antibody
1
Analyzing β-catenin Expression in Dct-Lac-Z Transgenic Mice
2
Immunohistochemical Analysis of β-Catenin and BDNF
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At 7 days after the operation, rats were anesthetized with 10% chloral hydrate (0.33 mL/kg) and then perfused with 4% paraformaldehyde. In brief, spinal cord tissues (T8/9) 3 mm rostral to the epicenter were harvested from the rats and fixed in 4% paraformaldehyde. Tissues were immersed in 30% sucrose overnight and sliced into cross-sections at a thickness of 5 μm. Sections were placed into a box with 0.01 M citric acid for antigen retrieval. Then, the sections were blocked using blocking buffer (5% normal goat serum and 0.1% Triton-X100 in PBS) at 4°C for 1 hour and incubated with primary antibodies (rabbit anti-β-catenin polyclonal antibody,1:400, Abcam; rabbit anti-BDNF polyclonal antibody, 1:500, Novus Biologicals, Littleton, CO, USA) at 4°C overnight. Next, the tissues were incubated with FITC-goat anti-rabbit IgG (1:400, Abcam) and the nuclei were stained with DAPI (1:1,000) solution. All slides were observed under a fluorescence microscope (Leica, Heidelberger, Germany) after being mounted with Permount™ Mounting Medium (Sigma-Aldrich, St. Louis, MO, USA). The optical density of fluorescence was analyzed using ImageJ2x software.
3
Skin Protein Extraction and Western Blot
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Proteins were extracted from the skin specimens by using of RIPA lysis buffer (Beyontime, China). The extracts were vortexed, centrifuged at 12000×g for 10min. The concentration was determined by using a BCA protein concentration determination kit (Beyontime, China). Then the extracts were added with the loading buffer and incubated in boiling water for 5 min, followed by electrophoresis (concentrating gel 80V, 30 min; separating gel 100V, 90 min). The proteins were transferred to a PVDF membrane (250 mA, 2 h), which was treated with methanol for 1 min and electrotransfer buffer for 15 min before use. After wash with TBST (5min × 5), the PVDF membrane was blocked with 5% defatted milk (2 h), incubated with rabbit anti-β-catenin polyclonal antibody (1:1000; Abcam, USA) at 4°C for overnight, and secondary antibody (1:10000; Boster, China) at 37°C for 1 h, then developed with chemoluminescence.
4
Western Blot Analysis of Spinal Cord Injury
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The rats were severally narcotized with 10% chloral hydrate (0.33 mL/kg) via intraperitoneal injection at 3 and 5 days post-surgery, and the T8–11 spinal cord (2 mm cephalad and caudally from the epicenter) was dissected out. The tissues were homogenized in RIPA lysis buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% NP-40, 0.25% Na-deoxycholate, 1 mM EDTA). Then, the concentrations of proteins were assessed using the bicinchoninic acid kit, adjusted to 2 μg/μL, and 40 μg of protein was resolved on 12% Tris-glycine SDS-PAGE gels. Then, the proteins were transferred to polyvinylidene fluoride membranes and incubated with primary antibodies (rabbit anti-β-catenin polyclonal antibody, 1:1,000, Abcam, Cambridge, UK; rabbit anti-caspase-3 polyclonal antibody, 1:1,000, Abcam; mouse anti-β-actin polyclonal antibody, 1:1,000, Abcam) at 4°C overnight after being sealed. On the following day, the membranes were washed three times with Tris-buffered saline (TBS; 150 mM NaCl, 100 mM Tris-HCl, pH 7.4) containing 0.1% Tween-20, and then incubated with goat anti-rabbit/mouse IgG (1:3,000, Abcam) at room temperature for 2 hours. Finally, the membranes were developed using a ChemiDoc-It™ TS2 Imager (UVP, LLC, Upland, CA, USA) and relative optical density was measured using ImageJ2x software (National Institutes of Health, Bethesda, MD, USA).
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