OP and OC were purchased from Sequoia Research Products Ltd. (Pangbourne, UK) and MedChemexpress Co., Ltd. (Monmouth Junction, NJ, USA), respectively. Maoto (lot: F47711, J04981) and kakkonto (lot: F25972, J26252) extract granules were purchased from Tsumura Co., Ltd. (Tokyo, Japan). EB and LPS (derived from the Salmonella enteric serotype Typhimurium) were obtained from Sigma Chemical Co., Ltd. (St. Louis, MO, USA). The rabbit anti-β-actin polyclonal antibody (43 kDa), rat anti-MRP4 monoclonal antibody (150 kDa), and horseradish peroxidase- (HRP-) conjugated goat anti-rat immunoglobulin (IgG) were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). The rabbit anti-OAT3 polyclonal antibody (59 kDa) was purchased from Biorbyt Ltd. (Cambridge, UK). HRP-conjugated goat anti-rabbit IgG was purchased from Enzo Life Sciences Inc. (Farmingdale, NY, USA). All other chemicals and solvents were of reagent or high-performance liquid chromatography (HPLC) grade and were used without further purification.
Rabbit anti β actin polyclonal antibody
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Rabbit anti-β-actin polyclonal antibody is a primary antibody that recognizes the β-actin protein, which is a widely expressed cytoskeletal protein involved in various cellular processes. This antibody is produced in rabbits and can be used for the detection and quantification of β-actin in a variety of applications.
Automatically generated - may contain errors
Lab products found in correlation
Ne per nuclear and cytoplasmic extraction reagent, by Thermo Fisher Scientific (2 mentions)
Hrp conjugated goat anti rabbit igg, by Enzo Life Sciences (1 mentions)
Novolin r, by Novo Nordisk (1 mentions)
Super ecl plus detection reagent, by Applygen (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Mouse monoclonal anti p53 antibody, by Cell Signaling Technology (1 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium dmem, by Merck Group (1 mentions)
Hepatocyte medium, by ScienCell (1 mentions)
Huh7 cells, by National Collection of Authenticated Cell Cultures (1 mentions)
Pvdf membrane, by Beyotime (1 mentions)
Horseradish peroxidase conjugated goat anti rabbit secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Rabbit anti mmp 2 polyclonal antibody, by Abcam (1 mentions)
Rabbit anti mmp 9 polyclonal antibody, by Abcam (1 mentions)
Radioimmunoprecipitation assay buffer, by Beyotime (1 mentions)
Enhanced chemiluminescence kit, by Beyotime (1 mentions)
Human recombinant tnfα, by R&D Systems (1 mentions)
Rabbit monoclonal anti egfr antibody d38b1, by Cell Signaling Technology (1 mentions)
Ag1478, by Cell Signaling Technology (1 mentions)
Normal armenian hamster igg, by Santa Cruz Biotechnology (1 mentions)
9 protocols using rabbit anti β actin polyclonal antibody
1
Characterization of Transporter Proteins
2
Affinity Purification of Anti-PPARγ Antibody
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Affinity-purified rabbit anti-PPARγ polyclonal antibody was purchased from Abcam Inc. (Cambridge, MA, USA; cat. no. ab19481). Human insulin (Novolin™ R) was from Novo Nordisk (Copenhagen, Denmark). Candesartan cilexetil was provided by Takeda Pharmaceutical Co., Ltd. (Osaka, Japan). Rabbit anti-β-actin polyclonal antibody was from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA; cat. no. sc-1616). Peroxidase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) was obtained from Beijing Zhongshan Golden Bridge Biotechnology Co., Ltd. (Beijing, China; cat. no. ZB-2301).
3
Quantifying PPAR-gamma and NF-kB in Ischemic Cortex
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Protein from cerebral ischemic cortex or cortical neurons was collected and diluted to a concentration of 0.5 mg protein/mL for the measurement of PPARγ and NF-κB 65. Equal amounts of protein were electrophoresed through a reducing sodium dodecyl sulfate polyacrylamide gel and electroblotted onto a polyvinylidene difluoride membrane. After incubation in blocking buffer (5% nonfat dried milk, phosphate-buffered saline, and 0.1% Tween-20) at room temperature for 1 hour, membranes were incubated with mouse anti-rat PPARγ monoclonal antibody (1 : 500; Santa Cruz Biotechnology, Santa Cruz, CA, USA), mouse anti-rat NF-κB 65 monoclonal antibody (1 : 500; Santa Cruz Biotechnology), or rabbit anti-β-actin polyclonal antibody (1 : 3,000; Santa Cruz Biotechnology) overnight at 4°C. Protein levels were detected with goat anti-rabbit or anti-mouse IgG-horseradish peroxidase-linked secondary antibodies (1 : 2,000; Santa Cruz Biotechnology) at room temperature for 1 hour and developed with Super ECL Plus detection reagent (Applygen Technologies, Beijing, China). Films were scanned and the intensities of immunoblot bands were quantified by densitometry using image analysis software (Image Master Total Lab version 1.00; Amersham Pharmacia Biotech, Tokyo, Japan). The band absorbance values were calculated as a ratio of PPARγ/β-actin or NF-κB 65/β-actin.
4
Cultivation and Characterization of Hepatic Cell Lines
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Huh-7 cells were purchased from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China) and maintained in Dulbecco’s modified Eagle’s medium (DMEM, Sigma) supplemented with 10% fetal bovine serum (Gibco, United States). HepAD38 and HepG2-NTCP cells were obtained from Xiamen University, HepAD38 cells were maintained in DMEM supplemented with 10% FBS and 400 μg/ml G418, and HepG2-NTCP cells were maintained in DMEM supplemented with 10% FBS, 2 μg/ml puromycin and 2 μg/ml Dox. PHHs were purchased from ScienCell and were maintained in hepatocyte medium (ScienCell, Carlsbad, CA, United States). All cells were incubated in a humidified atmosphere at 37°C and 5% CO2. Transfection was performed using Lipofectamine™ 3,000 (Invitrogen, United States) according to the manufacturer’s protocol. Rabbit anti-β-actin polyclonal antibody (100 μg/ml; sc-1, 616-R) and mouse anti-SIRT2 monoclonal antibody (200 μg/ml; sc-28, 298) were purchased from Santa Cruz Biotechnology (Dallas, TX, United States), and mouse anti-p53 monoclonal antibody (72 μg/ml; #48818) was obtained from Cell Signaling Technology (United States). The Human Sirtuin 2 (SIRT2) ELISA Kit was obtained from Abbexa Ltd. (abx383209; Cambridge, United Kingdom).
5
Western Blot Analysis of Bone-Related Proteins
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Total protein of ECs and SMCs were extracted in the radio immunoprecipitation assay buffer (Beyotime, China). After centrifuging, supernatants were collected for the determination of protein concentrations. Equal protein (50 μg) was loaded onto 10% SDS -polyacrylamide gels and transferred to PVDF membranes (Beyotime, China). The membranes were blocked with 5% bovine serum albumin in TBST at room temperature for 1 h, then incubated with primary antibodies: rabbit anti-BMP2 polyclonal antibody (Abcam, Boston, Massachusetts, USA; dilution 1:300), or anti-Runx2 polyclonal antibody (Abcam, USA; dilution 1:500), or rabbit anti-Msx2 polyclonal antibody (Santa Cruz Biotechnology, California, USA, dilution 1:500), or rabbit anti-Osterix polyclonal antibody (Abcam, USA; dilution 1:300), or rabbit anti-MMP-2 polyclonal antibody (Abcam, USA; dilution 1:500), or rabbit anti-MMP-9 polyclonal antibody (Abcam, USA; dilution 1:500), or rabbit anti-β-actin polyclonal antibody (Santa Cruz, USA, dilution 1:500 ) overnight at 4°C. After washed with TBST three times, membranes were incubated with horseradish peroxidase conjugated goatanti-rabbit secondary antibody (Santa Cruz, dilution 1:1000) for 1 h at 37°C. Immunodetections were performed with the Enhanced Chemiluminescence Kit (Beyotime, China).
6
Immunoblotting Analysis of Receptor Tyrosine Kinases
7
Protein Expression Analysis by Western Blot
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Proteins (30μg) were separated by SDS-PAGE and transferred to nitrocellulose membranes. Nonspecific binding was blocked in 5% nonfat milk in Tris-buffered saline (TBS) + 0.1% Tween-20 for 1 h. The membranes were probed overnight with primary antibodies for rabbit anti-Mfn2 (Abcam, Cambridge, MA), rabbit polyclonal anti-Bcl-2 antibody (Cell Signaling, USA), rabbit polyclonal anti-Bax antibody (Cell Signaling, USA), rabbit polyclonal anti-caspase3 antibody (Cell Signaling, USA), rabbit polyclonal anti-caspase9 antibody (Cell Signaling, USA), rabbit polyclonal anti-MMP-2 antibody (Cell Signaling, USA), rabbit polyclonal anti-MMP-9 antibody (Cell Signaling, USA), rabbit polyclonal anti- integrin β1 antibody (Cell Signaling, USA) and rabbit polyclonal anti-β-actin antibody (Santa Cruz Biotech, CA) in TBS-T containing 1% nonfat milk at 4°C, washed three times with TBS-T for 10 min, and were probed with horseradish peroxidase-conjugated secondary antibodies for 1 h. The membranes were washed three times and exposed in a standard enhanced chemiluminescence reaction according to manufacturer’s instructions (Pierce, Rockford, IL.). The results were normalized to β-actin signal intensity.
8
Protein Extraction and Western Blot Analysis
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After treatment, cells were washed twice with cold PBS and then pelleted, and then cell lysis reagent (Sigma-Aldrich) was added to the cell pellet for protein extraction. Protein concentration was determined by bovine serum albumin (BSA) assay using BSA as the reference, and the absorbance of the reacted product was measured at wavelength of 540–595 nm. Samples were boiled for 3 min before loading onto 12% sodium dodecyl sulfate (SDS)-polyacrylamide gel. After electrophoresis, the gels were electro blotted onto a polyvinylidene fluoride (PVDF) membranes (Merck Millipore, Whitehouse Station, NJ, USA). The antibodies used were as follows: anti-Bcl-2 rabbit polyclonal antibody (Abcam, Cambridge, MA, USA), anti-Bax rabbit polyclonal antibody (Abcam), anti-β-actin rabbit polyclonal antibody (Santa Cruz Biotechnology Inc., Dallas, TX, USA), and anti-rabbit IgG HRP (Abcam). The blot was visualized with an enhanced chemiluminescence (ECL) kit (Merck Millipore) and exposed to ECL Hyperfilm.
9
Western Blot Analysis of Circadian Proteins
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RIPA plus proteinase inhibitors were used to extract proteins from cells or SCN tissue samples. Nuclear/cytoplasm fractionation was performed using NE-PER nuclear and cytoplasmic extraction reagents (Thermo, USA). The cell or tissue lysates were normalised by measuring the protein concentration. The proteins were subsequently separated via SDS-PAGE and transferred to a nitrocellulose filter membrane for antibody recognition. The following antibodies were used: anti-CLOCK antibody (CST, USA), anti-NPAS2 antibody (Santa Cruz Biotechnology, USA), anti-CRY2 antibody (Santa Cruz Biotechnology, USA), Lamin B1 polyclonal antibody (ImmunoWay, USA), anti-β-actin rabbit polyclonal antibody (Santa Cruz Biotechnology, USA) and anti-GAPDH monoclonal antibody (MBL, Japan). The antibodies for PER1, CRY1 and BMAL1 were purchased from Abcam, UK. Quantity One software was used for the quantitative analyses of the protein expression.
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