The HepG2 cells were obtained from American Type Culture Collection (ATCC®, CCL-136™). LO2 cells (normal human liver cells line) were provided from Cell Bank of the Chinese Academy of Sciences (Shanghai, People’s Republic of China). Dulbecco’s Modified Eagle’s Medium and fetal bovine serum purchased from Gibco were used for cell culture. PTX, AgNO3, vitamin C, branched PEI, propidium iodide (PI), 2′,7′-dichlorofluorescein diacetate, 4′6-diamidino-2-phenylindole (DAPI), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) were all obtained from Sigma. Capase-3, poly(ADP-ribose) polymerase (PARP), H2X, P-p53, P53, TAKT, T-p38, and β-actin monoclonal antibody were purchased from Cell Signaling Technology. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay kit, Annexin-V-FLUOS staining kit, caspase-3 activity assay kit, and BCA protein assay kit were acquired from Beyotime Institute of Biotechnology (Shanghai, People’s Republic of China).
β actin monoclonal antibody
Manufactured by Cell Signaling Technology
Sourced in United States
The β-actin monoclonal antibody is a laboratory reagent used to detect the presence and abundance of the β-actin protein in biological samples. β-actin is a highly conserved cytoskeletal protein found in all eukaryotic cells and is commonly used as a loading control in various experimental techniques, such as Western blotting and immunocytochemistry.
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Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
2 7 dichlorofluorescein diacetate, by Merck Group (1 mentions)
Capase 3, by Cell Signaling Technology (1 mentions)
Terminal deoxynucleotidyl transferase dutp nick end labeling tunel assay kit, by Beyotime (1 mentions)
Branched pei, by Merck Group (1 mentions)
T akt, by Cell Signaling Technology (1 mentions)
Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (1 mentions)
Propidium iodide, by Merck Group (1 mentions)
Agno3, by Merck Group (1 mentions)
Vitamin c, by Merck Group (1 mentions)
Bca protein assay kit, by Beyotime (1 mentions)
Caspase 3 activity assay kit, by Beyotime (1 mentions)
Atcc ccl 136, by American Type Culture Collection (1 mentions)
Erk1 2 monoclonal antibody, by Cell Signaling Technology (1 mentions)
T gsh gssg, by Nanjing Jiancheng (1 mentions)
Goat anti mouse igg, by Cell Signaling Technology (1 mentions)
Goat anti rabbit igg, by Cell Signaling Technology (1 mentions)
Bullet blocking one, by Nacalai Tesque (1 mentions)
Transblot turbo mini size pvdf membrane, by Bio-Rad (1 mentions)
Chemidoc mp image lab, by Bio-Rad (1 mentions)
4 protocols using β actin monoclonal antibody
1
Evaluating Cytotoxicity of Nanoparticles
2
Biochemical Analyses of Oxidative Stress
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Assay kits for measuring superoxide dismutase (SOD), malondialdehyde (MDA), total glutathione/oxidized glutathione (T-GSH/GSSG), nitric oxide (NO), and total antioxidant capacity (T-AOC) were purchased from Nanjing Jiancheng Institute of Biotechnology (Nanjing, China). Anti-phospho-FGFR1 (Tyr653+Tyr654) polyclonal antibody (rabbit) was purchased from Bioss (Beijing, China). BDNF polyclonal antibody (rabbit) was purchased from Cloud-Clone Corp (Wuhan, Beijing). Goat-anti mouse IgG, goat-anti rabbit IgG, β-actin monoclonal antibody (rabbit), AKT (pan) monoclonal antibody (mouse), phospho-AKT (Ser473) monoclonal antibody (rabbit), phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) monoclonal antibody (rabbit), Erk1/2 monoclonal antibody (mouse), Bcl-2 monoclonal antibody (rabbit), and caspase-3 polyclonal antibody (rabbit) were purchased from Cell Signaling Technology (Danvers, United States). FGF-2 was produced in the EK’s lab, and the purification data, western blot data and activity data were presented in Supplementary Figure 1 .
3
Western Blot Protein Detection Protocol
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We used an SDS buffer (25% of 0.125M Tris-HCl (pH 6.8), 20% of glycerol, 4% of SDS, and 10% of 2-mercaptoethanol, with bromophenol blue included) to pellet cell pellets, extract proteins, and degrade DNA by sonication in ice-cold conditions. The protein extracts were separated in Mini-PROTEAN TGX Stain-Free gels (BIO RAD, CA, USA) by electrophoresis (60 mA, 40 min) and transferred to TransBlot Turbo Mini-size PVDF membranes (BIO RAD). The membranes were blocked with Bullet Blocking One (Nacalai Tesque, Kyoto, Japan). The antibodies we used were as follows: primary antibodies—SHC4 polyclonal antibody (rabbit, 75 kDa) (Sigma-Aldrich, Mizulli, USA) and β-actin monoclonal antibody (rabbit; 45 kDa) (Cell signaling technology, #4790, MA, USA) diluted 1000×, reacted at 4 °C in overnight. The membranes were washed with TBS-T, and the horseradish peroxidase (HRP)-conjugated anti-rabbit antibody (GE Healthcare, Chicago, IL, USA) diluted 5000× was reacted with membrane for 1 h. The membranes were washed with TBS-T three times before being exposed to the ClarityTM Western ECL Substrate (Bio Rad Laboratories, Hercules, CA, USA) as a chromogenic substrate for HRP. The protein bands were detected using a ChemiDoc MP ImageLab (Bio Rad Laboratories, Hercules, CA, USA).
4
Western Blot Analysis of Cellular Proteins
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The cell culture samples were lysed in modified Radioimmune Precipitation Assay (RIPA) Buffer (Applygen) containing a protease inhibitor cocktail (Roche). The protein concentrations were measured using the BCA assay kit (Thermo Fisher Scientific). A total protein of 10–20 μg from each sample were loaded and subjected to 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and then transferred to PVDF filters (Millipore). Immunoprobing of target proteins were performed by incubating overnight at 4 °C with the primary antibodies against p21 (#10355–1-AP, Proteintech), TCAB1(#14761–1-AP, Proteintech) or ubiquitin (#D058-3, MBL). After rinses, HRP-conjugated secondary antibodies used for western blotting analysis were as follows: anti-rabbit IgG (#3012; Signalway Antibody) and mouse IgG (#3032; Signalway Antibody). The even loading of isolated proteins was verified with β-actin monoclonal antibody (#3700S, Cell Signaling Technology) or GAPDH monoclonal antibody (#10494–1-AP, Proteintech). The blotted filters were visualized for signals using enhanced chemiluminescence (ECL) reagents (GE Healthcare) according to the manufacturer’s instructions.
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