Total proteins from MSB1, MSB1-SiRNA-3, and MSB1-SiRNA-NC cells were prepared with RIPA lysis buffer (Beyotime Institute of Biotechnology, China). After protein quantitation by UV spectrophotometer assay, equal amounts of the proteins were separated with 12% SDS–PAGE and electro-transferred onto a 0.2 micron nitrocellulose membrane. The blots were then blocked with 5% skim milk in PBST (phosphate buffered saline, pH 7.4, containing 0.05% Tween 20) for 1 h at room temperature, after which they were incubated with a rabbit polyclonal antibody against chPrPC (1 : 500, in PBST with 5% skim milk; preparation and preservation in our laboratory [13 ]) or a rabbit antibody against β-actin (1:1,000, to confirm equal sample loading; Sigma, USA) for 1 h at 37℃. After washing with PBST three times and incubating with HRP-conjugated goat anti-rabbit IgG (1: 5,000, in PBST with 5% skim milk; Sigma) for 1 h at 37℃, the membranes were washed three times in PBST, and the blots were stained with diaminobenzidine (DAB; PBST containing 0.5 mg/mL DAB, 0.3 g/mL mercuric chloride, and 0.33 µL/mL hydrogen peroxide). Band intensities were quantified using Image-Pro-Plus software and normalized to the quantity of β-actin.
Rabbit antibody against β actin
Manufactured by Merck Group
Sourced in United States
Rabbit antibody against β-actin is a laboratory reagent used to detect the presence and quantify the levels of the β-actin protein in biological samples. β-actin is a widely expressed and highly conserved cytoskeletal protein that plays a crucial role in various cellular processes. This antibody can be utilized in various immunoassay techniques, such as Western blotting, immunohistochemistry, and immunocytochemistry, to investigate the expression and distribution of β-actin in different cell types and tissues.
Automatically generated - may contain errors
Lab products found in correlation
Primary rabbit antibody against acetyl histone h4, by Cell Signaling Technology (2 mentions)
Ripa lysis buffer, by Santa Cruz Biotechnology (2 mentions)
Alkaline phosphatase conjugated goat anti rabbit igg antibody, by Merck Group (2 mentions)
Bradford assay, by Bio-Rad (2 mentions)
Hrp conjugated goat anti rabbit igg, by Merck Group (1 mentions)
Ripa lysis buffer, by Beyotime (1 mentions)
Ferric ammonium citrate, by Merck Group (1 mentions)
Cholesterol, by Dingguo (1 mentions)
Odyssey image studio, by LI COR (1 mentions)
Facsdiva software, by BD (1 mentions)
Odyssey fc imager, by LI COR (1 mentions)
Western blotting luminol reagent, by Santa Cruz Biotechnology (1 mentions)
Enhanced chemiluminescence, by Thermo Fisher Scientific (1 mentions)
Immobilon p polyvinylidene difluoride membrane, by Merck Group (1 mentions)
5 protocols using rabbit antibody against β actin
1
Western Blot Analysis of Prion Protein
2
Antibody-based Apoptosis Analysis Protocol
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Ferric ammonium citrate and a rabbit antibody against β-actin were obtained from Sigma-Aldrich (USA). Soya bean lecithin (a source of phospholipids) was purchased from Beijing Huaqing Meiheng Biotechnology Co. Ltd. (China). Cholesterol was purchased from Beijing Dingguo Biotechnology Co. Ltd. (China). A rabbit polyclonal antibody against L-ferritin was obtained from Abcam (United Kingdom), and rabbit monoclonal antibodies against B cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were purchased from Santa Cruz Biotechnology (USA). The terminal deoxynucleotidyl transferase-mediated FITC-dUDP nick-end labeling (TUNEL) kit was from TaKaRa (Japan). All other analytical reagents and chemicals were purchased from Bio-High Technology Development Co. Ltd. (China).
3
Dual Detection of GFP and Viral Proteins
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GFP-positive cells were detected using multi-parameter fluorescent activated cell analysis (FACs) (Facs Canto-Beckton Dickinson). Intracellular staining of Zta was undertaken with the mouse monoclonal antibody BZ1 (33 (link)), with anti-mouse IgG coupled to Donkey F(ab’)2 as a secondary reagent (anti-mouse IgG -Alexa Fluor® 647) and the FIX & PERM® Cell Permeabilization Kit (LifeTechologies). Intracellular staining of VCA was undertaken with the mouse monoclonal antibody VCA-gp125 (clone L2 MAB8184, Millipore) and the same secondary reagent and conditions. Dual parameters of GFP and Alexa-fluor 647-coupled staining were collected and the double positive cells identified using BD FACSDiva™ Software (Beckton Dickinson).
Cells were harvested in sample buffer (4% SDS, 20% glycerol, 10% 2-mercaptoethanol, 0.004% bromophenol blue and 0.125 M Tris–HCl, pH 6.8.) and total proteins were fractionated by SDS-PAGE in a 12% gel (Novex). After transfer to nitrocellulose membrane, Zta and actin were detected by immunoblotting with the BZ1 mouse monoclonal antibody against Zta (33 (link)) and a rabbit antibody against β-actin (Sigma). Species-specific IR-labeled anti-mouse and anti-rabbit antibodies (Licor) were used in the second layer and the resulting signal was detected on an Odyssey Fc Imager and quantified using Odyssey Image Studio (Licor).
Cells were harvested in sample buffer (4% SDS, 20% glycerol, 10% 2-mercaptoethanol, 0.004% bromophenol blue and 0.125 M Tris–HCl, pH 6.8.) and total proteins were fractionated by SDS-PAGE in a 12% gel (Novex). After transfer to nitrocellulose membrane, Zta and actin were detected by immunoblotting with the BZ1 mouse monoclonal antibody against Zta (33 (link)) and a rabbit antibody against β-actin (Sigma). Species-specific IR-labeled anti-mouse and anti-rabbit antibodies (Licor) were used in the second layer and the resulting signal was detected on an Odyssey Fc Imager and quantified using Odyssey Image Studio (Licor).
4
Histone H4 Acetylation Analysis
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Chicken HD11 cells were stimulated with 0.2M lactose in the presence or absence of 2mM butyrate for 6, 12, or 24h, followed by wash with phosphate buffered saline and lysis in the radioimmunoprecipitation (RIPA) lysis buffer (Santa Cruz Biotechnology). Protein concentration was measured using the Bradford Assay (Bio-Rad). To determine the levels of histone H4 acetylation, 20μg proteins were separated in 12.5% SDS-PAGE gels and then transferred to polyvinylidene difluoride (PVDF) membranes. After overnight blocking in the blocking buffer containing 5% dry skim milk in TTBS (0.05% Tween 20, 20mm Tris-HCl, 150mm NaCl, pH 7.5) at 4°C, the membranes were incubated with a primary rabbit antibody against acetyl-histone H4 (Cell Signaling, Danvers, MA, United States) or a rabbit antibody against β-actin (MilliporeSigma) in the blocking buffer for 1h at room temperature. After three washes in TTBS, the membrane was incubated with an alkaline phosphatase-conjugated goat anti-rabbit IgG antibody (MilliporeSigma) for 45min at room temperature. Western blots were visualized with Western Blotting Luminol Reagent (Santa Cruz Biotechnology).
5
Acetyl-histone H4 Quantification in Chicken HTC Cells
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Chicken HTC cells were treated with 2 mM butyrate, 20 μM quercetin, or their combination for 6 or 12 h, followed by lysis in the radioimmunoprecipitation (RIPA) lysis buffer (Santa Cruz Biotechnology). The resulting proteins were quantified using the Bradford Assay (Bio-Rad), followed by loading of 20 µg proteins from each sample in 12.5% SDS-PAGE gels and transferring to Immobilon-P® polyvinylidene difluoride membranes (MilliporeSigma). After overnight blocking in 5% dry skim milk in TTBS (0.05% Tween 20, 20 mM Tris-HCl, 150 mM NaCl, pH 7.5) at 4°C, the membranes were incubated with a primary rabbit antibody against acetyl-histone H4 (Cell Signaling, Danvers, MA; diluted 1:500) or a rabbit antibody against β-Actin (MilliporeSigma; diluted 1:1000) in the blocking buffer for 1 h at room temperature. After three washes in TTBS, the membranes were incubated with an alkaline phosphatase-conjugated goat anti-rabbit IgG antibody (MilliporeSigma; diluted 1:2,000) for 45 min at room temperature, followed by visualization using enhanced chemiluminescence (ThermoFisher Scientific). The band intensity of acetyl-histone H4 was quantified as the area under the curve using ImageJ (https://imagej.nih.gov/ij/ ) and further normalized against the band intensity of β-Actin for each sample.
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