Paclitaxel and SR13800 were purchased from Sigma-Aldrich (St. Louis, MO, USA). Antibodies against β-actin (#4970) and LDHA (#3582) were purchased from Cell Signaling (Beverly, MA, USA). Monoclonal mouse antibody against MCT1 was purchased from ThermoFisher Scientific (#MA5-18288, Waltham, MA, USA). Control siRNA and MCT1 siRNA (5′-GAGGAUGAAUCACUAAGUA-3′) were synthesized by Ribobio (Guangzhou, China).
Antibodies against β actin
Manufactured by Cell Signaling Technology
Sourced in United States
Antibodies against β-actin are a type of laboratory equipment used to detect and quantify the expression of the β-actin protein in biological samples. β-actin is a widely expressed cytoskeletal protein that is commonly used as a reference or housekeeping gene for normalization in various experimental techniques, such as Western blotting, immunohistochemistry, and flow cytometry.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Merck Group (3 mentions)
Gapdh, by Cell Signaling Technology (3 mentions)
P jnk, by Cell Signaling Technology (3 mentions)
Lipopolysaccharides (lps), by Merck Group (3 mentions)
Phospho akt ser473, by Cell Signaling Technology (2 mentions)
Caspase 3, by Cell Signaling Technology (2 mentions)
Trizol reagent, by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Merck Group (2 mentions)
Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (2 mentions)
Dulbecco s modified eagle s medium dmem, by Nacalai Tesque (2 mentions)
P erk1 2, by Cell Signaling Technology (2 mentions)
P iκbα, by Cell Signaling Technology (2 mentions)
Dextran sulfate sodium (dss), by MP Biomedicals (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Cleaved caspase 3, by Cell Signaling Technology (2 mentions)
Phospho erk1 2 thr202 tyr204, by Cell Signaling Technology (2 mentions)
Paclitaxel, by Merck Group (1 mentions)
Aldh1a1, by Cell Signaling Technology (1 mentions)
Lamin a c, by Cell Signaling Technology (1 mentions)
P ampk, by Cell Signaling Technology (1 mentions)
34 protocols using antibodies against β actin
1
Paclitaxel and SR13800 Cytotoxicity Assay
2
Immunoblotting, IHC, and IF Protocol
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An Ab against SDPR (HPA039325 ; Merck, Darmstadt, Germany) was used for immunoblotting (1:250), immunohistochemistry (1:1000), and immunofluorescence (1:500) analyses. Antibodies for immunoblotting against N‐cadherin, vimentin, and snail (1:1000; EMT Antibody Sampler Kit, no. 9782), ALDH1A1 (1:1000; no. 54135), AKT (1:1000; no. 4691) and phospho‐AKT (Ser473; 1:2000; no. 4060) were obtained from Cell Signaling Technology (Danvers, MA, USA). An Ab against ILK1 (no. 3862; Cell Signaling Technology) was used for immunoblotting (1:1000) and immunofluorescence (1:100) analyses. Antibodies against β‐actin (1:1000; 13E5, HRP conjugate, no. 5125) and lamin A/C (1:1000; no. 2032), as immunoblotting loading controls, were also purchased from Cell Signaling Technology.
3
Western Blot Analysis of Cellular Proteins
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Cells were washed in PBS, and whole-cell extracts were prepared in lysis buffer (Tris-HCl (20 mM), pH 7.4, NaCl (150 mM), and glycerol (10% (v/v)), Nonidet P-40 (0.2%), EGTA (1 mM), EDTA (1 mM), PMSF (1 mM), NaF (10 mM), leupeptin (20 mM), aprotinin (5 mg/mL), and sodium orthovanadate (1 mM)) and centrifuged at 8,000 ×g for 15 min. Protein concentrations were measured using the BCA assay (Pierce). Protein (50 μg) was separated on a 15% (w/v) sodium dodecyl sulphate polyacrylamide gel and then transferred to PVDF membranes (Millipore, Billerica, MA, USA). Membranes were incubated overnight with primary antibodies specific to UCP2 (1 : 1000, Biolegend), p-AMPK (1 : 2000, Cell Signaling Technology), caspase-3 (1 : 1000, Cell Signaling Technology, Beverly MA, USA), and LC3 (1 : 2000, Cell Signaling Technology) at 4°C overnight. The positive reaction against these antibodies was visualized by enhanced chemiluminescence (ECL, Santa Cruz) reagent, followed by exposure to Kodak X-Omat X-ray film. After rinsing the membranes with acetonitrile for 10 min, the membranes were rehybridized with antibodies against β-actin (1 : 2000, Cell Signaling Technology) as the loading control. Relative density of protein bands was determined using ImageJ software (National Institutes of Health, USA).
4
Molecular Mechanisms of Vascular Remodeling
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Norepinephrine bitartrate monohydrate was obtained from MedChem Express (Monmouth Junction, NJ, USA). Propranolol hydrochloride and losartan were bought from Tocris Bioscience (Bristol, Avon, UK). Phentolamine hydrochloride and GW4869 were purchased from Sigma (St. Louis, MO, USA). GW4869 was dissolved in 1% dimethyl sulfoxide (DMSO). PBS containing 1% DMSO was used as the control. Other chemicals were dissolved in PBS.
Antibodies against α-SMA (1:5000), CD9 (1:1000), TSG101 (1:2000), calnexin (1:2000) and ACE (1:1000) were purchased from Abcam (Cambridge, MA, USA). Antibodies against PCNA (1:5000) and PECAM-1 (1:2000) were obtained from Protein Tech Group Inc. Antibody against CD63 (1:1000) was acquired from Wanleibio. (Shengyang, China). Antibody against vimentin (1:2000) was purchased from Abways Technology, Inc. (Shanghai, China). Antibodies against β-actin (1:10000) and GAPDH (1:10000) were bought from Cell Signaling Technology (Beverly, MA, USA). These antibodies were used for Western blot analyses. GAPDH and β-actin were used as loading control.
Antibodies against α-SMA (1:5000), CD9 (1:1000), TSG101 (1:2000), calnexin (1:2000) and ACE (1:1000) were purchased from Abcam (Cambridge, MA, USA). Antibodies against PCNA (1:5000) and PECAM-1 (1:2000) were obtained from Protein Tech Group Inc. Antibody against CD63 (1:1000) was acquired from Wanleibio. (Shengyang, China). Antibody against vimentin (1:2000) was purchased from Abways Technology, Inc. (Shanghai, China). Antibodies against β-actin (1:10000) and GAPDH (1:10000) were bought from Cell Signaling Technology (Beverly, MA, USA). These antibodies were used for Western blot analyses. GAPDH and β-actin were used as loading control.
5
Western Blot Analysis of Protein Expression
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Total protein was isolated from the cells using radioimmunoprecipitation assay (RIPA) buffer (Beyotime Biotechnology, China) that was supplemented with a protease inhibitor cocktail. Protein extracts were separated via 8%–12% SDS-polyacrylamide gel electrophoresis and transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Billerica, MA, USA). The membranes were then blocked in 5% fat-free milk and incubated with primary antibodies overnight at 4°C. Following washing with Tris-buffered saline in Tween 20 (TBST), the membranes were incubated with secondary antibodies at room temperature. Signal was visualized via enhanced chemiluminescence (ECL) using an ECL kit (Thermo Fisher Scientific, USA). Antibodies against β-actin (catalog #4970), ERCC1 (catalog #12345), and E2F1 (catalog #3742) were from Cell Signaling Technology (Beverly, MA, USA).
6
Evaluation of P53 and MDM2 in Cell Cycle
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Dulbecco's modified Eagle's medium (DMEM; #08458-45) was obtained from Nacalai Tesque (Kyoto, Japan). Fetal bovine serum (FBS; #172012) and paclitaxel (T7402) were purchased from Sigma Aldrich (St. Louis, MO). Antibodies against MDM2 (sc-965) and P53 (sc-126) were obtained from Santa Cruz Biotechnology (Dallas, TX). The cell cycle regulation antibody sampler kit (#9932), antibodies against β-actin (#4967), Rb-control (#9303), and phospho-Rb (#9307) were obtained from Cell Signaling (Danvers, MA). Lipofectamine 2000 Transfection Reagent (#11668027) and TRIzol Reagent (#15596-018) were from Life Technologies (Carlsbad, CA).
7
Cytokine and Antibody Stimulation Assay
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Recombinant mouse IFNγ, TNFα, IL-17A, and antibodies against IL-17A were from eBiosciences (La Jolla, CA, USA). Recombinant mouse IL-2 was from R&D Systems (Minneapolis, MN, USA). Antibodies against β-actin, GAPDH, iNOS, p65, p-IκBα, p-p65, p-JNK, and p-ERK1/2 were from Cell Signaling Technology (Danvers, MA, USA). Antibody against Act1 was from Santa Cruz Biotechnology (Dallas, TX, USA). L-NMMA, propidium iodide, and actinomycin D were purchased from Sigma-Aldrich (St Louis, MO, USA). Concanavaline A (ConA) was from Vector Labs (Burlingame, CA, USA).
C57BL/6 mice were purchased from the Shanghai Laboratory Animal Center of the Chinese Academy of Sciences (Shanghai, China) and maintained under specific pathogen-free conditions in the vivarium of Shanghai Jiao Tong University School of Medicine with water and food provided ad libitum. All animal protocols are approved by our Institutional Animal Care and Use Committee.
C57BL/6 mice were purchased from the Shanghai Laboratory Animal Center of the Chinese Academy of Sciences (Shanghai, China) and maintained under specific pathogen-free conditions in the vivarium of Shanghai Jiao Tong University School of Medicine with water and food provided ad libitum. All animal protocols are approved by our Institutional Animal Care and Use Committee.
8
Isolation and Characterization of Melianodiol
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Melianodiol (MN) was isolated from the fruits of M. azedarach by our research group. The structure of MN is shown in Fig. 1A (Xu et al., 2008 (link)). NO, IL-6, IL-10, and TNF-α ELISA kits were provided by Shanghai MLBIO Biotechnology Co. Ltd. (Shanghai, China). LPS was obtained from Sigma (Sigma Aldrich, St. Louis, MO, USA). 5-Aminosalicylic acid was provided by Shanghai Aladdin Biochemical Co. Ltd. (Shanghai, China). DSS was provided by MP Biomedicals (USA) and indomethacin by Aladdin (USA). Antibodies against β-actin, JAK2, STAT3, IKKα/β, NF-κBP65, IKBα, p-IKKα/β, p-IκBα, p-P65, iNOS, eNOS, Nrf-2, HO-1, and Keap1 were provided by Cell Signaling Technology (Danvers, MA, USA). SOD, NO, GSH, and malondialdehyde (MDA) were obtained from the Nanjing Jian-Cheng Bioengineering Institute (Jiangsu, China).
9
Synthesis and Evaluation of β-Carboline Derivatives
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Twelve C1-, C3-, or C6-modified β-carboline derivatives were synthesized as previously described [27 (link),28 (link)]. The structures of these β-carboline derivatives are summarized in Table 1 .
Dynasore, U0126, LY294002, and ribavirin were purchased from Sigma-Aldrich (St. Louis, MO, USA). All compounds were dissolved in dimethyl sulfoxide (DMSO) for in vitro studies. The protease inhibitor cocktail and the phosphatase inhibitor were obtained from TargetMol (Boston, MA, USA). Antibodies against β-actin, p-ERK, ERK, p-Akt, Akt, goat anti-rabbit IgG, and goat anti-mouse IgG were purchased from Cell Signaling Technology (Topsfield, MA, USA). The mAb against NDV nucleoprotein was prepared in our laboratory.
DF-1, Hela, Vero, and BHK-21 cells originally obtained from ATCC (Manassas, VA, USA) were purchased from the Cell Bank of Chinese Academy Sciences (Shanghai, China). These cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with 10% fetal bovine serum (Gibco, Amarillo, TX, USA) at 37 °C with 5% CO2.
Four NDV strains, including F48E9, Blackbird/China/08, PPMV-1/SX-01/Ch/15, and La Sota were provided by the College of Veterinary Medicine, Northwest A&F University (Xianyang, China). These viruses were plaque-purified three times. Viruses were propagated in 9-day-old to 11-day-old SPF chicken embryos, titrated by plaque assay, and stored at –80 °C.
Dynasore, U0126, LY294002, and ribavirin were purchased from Sigma-Aldrich (St. Louis, MO, USA). All compounds were dissolved in dimethyl sulfoxide (DMSO) for in vitro studies. The protease inhibitor cocktail and the phosphatase inhibitor were obtained from TargetMol (Boston, MA, USA). Antibodies against β-actin, p-ERK, ERK, p-Akt, Akt, goat anti-rabbit IgG, and goat anti-mouse IgG were purchased from Cell Signaling Technology (Topsfield, MA, USA). The mAb against NDV nucleoprotein was prepared in our laboratory.
DF-1, Hela, Vero, and BHK-21 cells originally obtained from ATCC (Manassas, VA, USA) were purchased from the Cell Bank of Chinese Academy Sciences (Shanghai, China). These cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with 10% fetal bovine serum (Gibco, Amarillo, TX, USA) at 37 °C with 5% CO2.
Four NDV strains, including F48E9, Blackbird/China/08, PPMV-1/SX-01/Ch/15, and La Sota were provided by the College of Veterinary Medicine, Northwest A&F University (Xianyang, China). These viruses were plaque-purified three times. Viruses were propagated in 9-day-old to 11-day-old SPF chicken embryos, titrated by plaque assay, and stored at –80 °C.
10
Western Blotting Protocol for Immune Markers
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Proteins were extracted using lysis buffer with protease and phosphatase inhibitors. After boiling with loading buffer for 10 min, protein separation occurred through SDS-PAGE, followed by transfer to PVDF membranes. After blocking with 5% nonfat milk for 1.5 h at room temperature, membranes were incubated with primary antibodies overnight at 4 °C. Antibodies against RORγ (#ab113434, 1:1000), IL-17A(#ab193955, 1:1000), and TFF3(#ab272927, 1:1000) were obtained from Abcam (Cambridge, USA). Antibodies against β-actin (#3700S, 1:1000) and GAPDH(#5174S, 1:1000) were obtained from Cell Signaling Technology™ (CST, Danvers, MA, USA). Then, HRP-conjugated secondary antibodies (CAT: 7074# and 7076#, 1:5000, CST, USA) were applied for 1 h at room temperature. Protein bands were detected and quantified using chemiluminescence and Image Lab software.
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