The A2 β-casein fraction was resolved on a 12.5% acrylamide gel at 20 mA for 1 h using a Mini-Protean® Tetra System and PowerPacTM HV (Bio-Rad, USA), according to the method of Laemmli (1970) (link). The gel was stained for 2 h with a coomassie blue solution (0.3 M coomassie blue G-250, 40% methanol, and 7% glacial acetic acid). Bands were analyzed using the Molecular Imager® GelDocTM XR plus Imaging system and the Image LabTM software 5.1 (Bio-Rad, USA).
Powerpac hv
Manufactured by Bio-Rad
Sourced in United States
The PowerPac HV is a high-voltage power supply for electrophoresis applications. It provides a consistent and reliable power source for various types of gel electrophoresis, including SDS-PAGE, agarose, and native gel systems. The device features adjustable voltage, current, and time settings to accommodate different experimental requirements.
Automatically generated - may contain errors
Lab products found in correlation
Mini protean tetra system, by Bio-Rad (4 mentions)
Molecular imager geldoc xr plus imaging system, by Bio-Rad (4 mentions)
Image labtm software 5, by Bio-Rad (2 mentions)
Gel doc uv system, by Bio-Rad (1 mentions)
Image labtm software version 5, by Bio-Rad (1 mentions)
Porcine plasma, by Merck Group (1 mentions)
Coomassie brilliant blue g 250, by Bio-Rad (1 mentions)
Porcine igg, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Pdquest 2 d analysis software, by Bio-Rad (1 mentions)
Versadoc 3000, by Bio-Rad (1 mentions)
Polyvinylidene fluoride (pvdf), by Thermo Fisher Scientific (1 mentions)
Ma0151, by Meilun (1 mentions)
Rs0002, by Immunoway (1 mentions)
8 protocols using powerpac hv
1
SDS-PAGE Analysis of A2 β-casein
2
Beef Hydrolysis Profiling by SDS-PAGE
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We performed SDS-PAGE using 12.5% acrylamide as described by Laemmli (1970) to evaluate the hydrolysis profile of beef samples. A 2× sample buffer (0.125 M Tris, 4% SDS, 20% glycerol, 2% β-mercaptoethanol, pH 6.8) was added to the supernatant of each sample in a 1:1 ratio, centrifuged for 5 min at 9,000×g, and heated at 95°C for 10 min. Electrophoresis was performed at 20 mA per gel for 1 h using PowerPacTM HV (BioRad) and Mini-Protean® Tetra System (BioRad). After electrophoresis, the gel was stained for 1 h with a coomassie blue solution (0.025% coomassie blue g-250, 40% methanol, and 7% glacial acetic acid) and analyzed using Molecular Imager® GelDocTM XR plus Imaging system (BioRad) and Image LabTM Software 5.1 (BioRad).
3
Spectrophotometric Analysis of Biomolecules
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Visual measurements were carried out on a portable spectrophotometer (NS810, Shenzhen 3nh Technology Co., Ltd). The electrophoresis system consisted of a DYCP-31E vertical electrophoresis tank (Beijing Liuyi Instrument Plant, China) and a steady voltage electrophoresis power supply (PowerPacTM HV, Bio-Rad, USA). Images were recorded by the Gel Doc UV system (Bio-Rad, USA).
4
SDS-PAGE Protein Separation and Analysis
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This procedure was carried out on a 12.5% acrylamide gel, as described by Laemmli (1970) (link). The sample was mixed with 2× sample buffer (0.125 M Tris-HCl pH 6.8, 4% SDS, 20% glycerol, and 2% β-mercaptoethanol) and then heated at 95℃ for 10 min. Electrophoresis was performed at 20 mA for 1 h, using a Mini-Protean® Tetra System and PowerPacTM HV (Bio-Rad, Hercules, USA). The gel was stained for 1 h with a Coomassie blue solution (0.025% Coomassie blue g-250, 40% methanol, and 7% glacial acetic acid). Analysis of the bands on the gel was performed using a Molecular Imager® GelDocTM XR plus Imaging system and the Image LabTM software version 5.1 (Bio-Rad).
5
SDS-PAGE Analysis of Purified Protein Complex
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The protein composition of the purified PIC was analyzed by SDS-PAGE in a 12.5% acrylamide gel as described by Laemmli (1970) (link). The samples, which included porcine IgG (Sigma), porcine plasma, PIC, and bovine serum albumin (BSA, Sigma), were mixed with sample buffer (0.125 M Tris-HCl, 4% SDS, 20% glycerol, and 2% β-mercaptoethanol, pH 6.8) and heated for 15 min at 98°C. Electrophoresis was conducted at 20 mA (per gel) for 1 h using a Mini-Protean® Tetra System and PowerPacTM HV (Bio-Rad, USA), and the gel was stained with Coomassie® brilliant blue G-250 (Bio-Rad) for 2 h. The bands on the gel were analyzed with a Molecular Imager® GelDocTM XR plus Imaging system and Image LabTM software (version 5.1; Bio-Rad).
6
Gastric Proteome Analysis by 2D-PAGE
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The stomach total protein (850 µg) was loaded into a commercial prefabricated IPG strip, which was rehydrated for 13 h at 50 V and 20°C (non-linear, 17 cm; pH 3–10; Bio-Rad Laboratories, Inc.). Isoelectric focusing was performed with a voltage gradient of 250, 500 and 2,000 V for 1 h each, and 8,000 V for 3 h, and then run at 8,000 V to a total of ~60 kVh. Prior to performing SDS-PAGE, the focused strips were equilibrated in 1% (w/v) DTT in equilibration buffer for 15 min, and in alkylating equilibration buffer for another 15 min, which contained 1% (w/v) iodoacetamide. Using a PowerPac HV, the strips were sealed on a 12.5% SDS-PAGE gel for electrophoresis (Bio-Rad Laboratories, Inc.). The gel, visualized using 0.08% CBB G-250, was digitized through a high-precision scanner (Versa Doc 3,000; Bio-Rad Laboratories, Inc.). Using PDQuest 2-D analysis software, protein spot assessment, measuring and matching were performed (version 8.0; Bio-Rad Laboratories, Inc.). The protein spots were automatically matched and carefully edited. The differentially expressed proteins were considered to be spots showing significant differences in intensity and present on at least two of the three gels in a single treatment (P≤0.05).
7
C4BPA Knockdown and Overexpression Analysis
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Cells transfected with pGPU6-C4BPA-shRNA1 and pBI-CMV3-C4BPA were harvested 24 h after transfection. After 24 h of transfection, trypsinize the cells and wash them three times with PBS. The cells were then resuspended in RIPA buffer with protease inhibitors (MA0151, Meilunbio). The cell lysate was removed by centrifugation at 12,000 rpm/min for 10 min at 4°C, and the supernatant was collected. The BCA protein quantitative analysis method determines the sample’s total protein concentration. For Western blot (PowerPac™ HV, Bio-Rad), separate the same amount of protein by SDS-PAGE and then transfer it to PVDF (88518, Thermo-Fisher) membrane with a membrane transfer machine. After blocking with 1×TBS buffer containing 1% BSA for 2 h, wash the PVDF membrane with TBST (B040126, Sangon Biotech, China) solution. Then, the membrane was incubated with a primary antibody at 4°C for 12 h. Incubate the membrane with a secondary antibody (RS0002, Immunoway) (diluted in 1% BSA) for 2 h. The enhanced chemiluminescence HRP substrate was used to observe protein bands. After taking a photo with a chemiluminescence instrument, grayscale analysis software (Tanon) was used to calculate the relative expression of the target protein in different samples. Primary antibodies were purchased from Bioss, China.
8
Electrophoretic Deposition of Iron Oxide Nanoparticles
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For electrophoretic deposition, an iron oxide nanoparticle suspension was prepared by adding the iron oxide nanoparticle stock suspension at either one or 5 vol% to a 0.01 M hydrochloric acid (HCl) in isopropanol (IPA). This EPD suspension yielded an effective pH (pHe) of 2.3, providing the particles with a highly positive surface charge (approximately +30 mV). In the EPD bath a graphite block (40.1 × 26.1 × 3.3 mm) was used as the positively charged electrode, and was placed parallel and 1.5 cm away from the substrate. A power supply (Bio-Rad PowerPac HV) was used to generate an electric field between the two electrodes, with the positive electrode connected to the graphite block (counter electrode) and the negative electrode connected to the substrate, resulting in the particles moving towards and depositing on the substrate. The specific field and time deposition conditions are detailed in Table 1 . After EPD, the deposited films were allowed to dry in air overnight.
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