SW-60, Lovo, HCT-116, NCM460, and HEK293 cells were grown and routinely maintained in Dulbecco modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 100 U/mL penicillin, and 100 mg/mL streptomycin. Cells were cultured at 37°Cin 5% CO2 and 95% humidified air. For establishment of stable MAGP2 overexpressing cells, overexpressing plasmid was constructed by Inovogen, China; then, the plasmid was packed by a lentivirus system (Inovogen, China). After removing the medium of the cells, the lentiviral supernatant was added (with 6 μg/mL polybrene). After 1 day, the supernatant was removed, and the cells were screened for 2 weeks using 10 μg/mL puromycin (Sigma-Aldrich, Shanghai, China). Anti-MAGP2 antibodies were purchased from Abcam (ab171737; Shanghai, China); anti-β-actin antibodies were purchased from Affinity Biosciences (AF7018; Guangzhou, China).
Anti β actin antibody
Manufactured by Affinity Biosciences
Sourced in United States, China
The Anti-β-actin antibody is a primary antibody that specifically binds to the β-actin protein. β-actin is a ubiquitously expressed cytoskeletal protein that plays a fundamental role in various cellular processes. This antibody can be used for the detection and quantification of β-actin in various experimental techniques, such as Western blotting, immunohistochemistry, and immunocytochemistry.
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Lab products found in correlation
Pvdf membrane, by Merck Group (12 mentions)
Bca protein assay kit, by Bio-Rad (2 mentions)
Antibody against ldha, by Affinity Biosciences (2 mentions)
Puromycin, by Merck Group (1 mentions)
Annexin 5 fitc apoptosis detection kit, by Keygen Biotech (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Trizol reagent, by Takara Bio (1 mentions)
Anti sirt1 antibody, by Cell Signaling Technology (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
Ros detection kit, by BestBio (1 mentions)
Anti bcl 2, by Affinity Biosciences (1 mentions)
Anti bax, by Affinity Biosciences (1 mentions)
Goat anti rabbit secondary antibody, by Affinity Biosciences (1 mentions)
Y 27632, by Merck Group (1 mentions)
Ccg 1423, by Apexbio (1 mentions)
Fura 2 acetoxymethylester fura 2 am, by Dojindo Laboratories (1 mentions)
Anti α smaantibody, by Affinity Biosciences (1 mentions)
Anti rhoa antibody, by Abcam (1 mentions)
Horseradish peroxidase hrp labeled secondary antibody, by Santa Cruz Biotechnology (1 mentions)
21 protocols using anti β actin antibody
1
MAGP2 Overexpression Cell Lines
2
Protein Extraction and Western Blot Analysis
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Total protein was extracted from RCSECs and rat choroid tissues. The samples were gently washed with PBS three times and then incubated for 15 min in lysis buffer. Total proteins were collected by centrifugation at 12,000×g for 10 min, and the precipitates were discarded. The proteins (20 µg) were separated by SDS-polyacrylamide gel electrophoresis and transferred to PVDF membranes. The nonspecific binding sites were blocked with TBST supplemented with 10% skim milk powder at 37 °C for 1 h, and then the membranes were incubated with specific antibodies at 4 °C overnight. Anti-β-actin antibodies (1:10,000, Affinity, China) were used as the internal control. After incubation with a secondary antibody, an enhanced chemiluminescent reagent (BioTech, Beijing, China) was used to detect the antibodies.
3
Nimodipine Cytoprotection against Oxidative Stress
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CG was from Chengdu Keloma Biotechnology Co., Ltd. (Chengdu, China); the purity was greater than 98%. Nimodipine was purchased from Guangdong Huanan Pharmaceutical Group Co., Ltd. (Guangdong, China). Dulbecco's modified Eagle's medium (DMEM), glucose-free DMEM, fetal bovine serum (FBS), and penicillin-streptomycin were obtained from Gibco (Grand Island, NY, USA). Cell Counting Kit-8 (CCK-8) was obtained from Dojindo Laboratories (Kumamoto, Japan). Lactate dehydrogenase (LDH), superoxide dismutase (SOD), and malondialdehyde (MDA) assay kits were obtained from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). The Annexin V-FITC Apoptosis Detection Kit was from KeyGEN BioTECH (Nanjing, China). The reactive oxygen species (ROS) Detection Kit was obtained from BestBio (Shanghai, China). PGC-1α enzyme-linked immunosorbent assay (ELISA) kits were from Jiangsu Enzyme Biotechnology Co., Ltd. (Jiangsu, China). Anti-SIRT1 antibody was supplied by Cell Signaling Technology (Danvers, MA, USA). Anti-FOXO1, anti-Bcl-2, anti-Bax, and anti-β-actin antibodies, and goat anti-rabbit secondary antibody were purchased from Affinity (Cincinnati, OH, USA). The TRIzol reagent was supplied by Takara Biomedical Technology (Shiga, Japan). The reverse-transcribed cDNA synthesis kit and qPCR kit were from Beijing TsingKe Biotech Co., Ltd. (Beijing, China).
4
Vascular Smooth Muscle Regulation Assays
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NaHS,
ACh, Atr, C3TF, PPG, and Y27632 were
purchased from Sigma Chemical (St. Louis); Asp was purchased from
Solarbio (Beijing, China); KD025 was purchased from Merck (Darmstadt,
Germany); CCG-1423 was purchased from ApexBio (Houston); fura-2-acetoxymethyl
ester (Fura-2 AM) was purchased from Dojindo (Kumamoto, Japan); anti-α-SMA
antibody, anti-β-actin antibody, and anti-p-MLC antibody were
purchased from Affinity Biosciences (Changzhou, China); anti-CSE antibody
and anti-RhoA antibody were purchased from Abcam (San Francisco);
anti-3-MST antibody was purchased from Santa Cruz Biotechnology (Santa
Cruz); anti-factor VIII antibody was purchased from Shanghai Fushen
Biotechnology Co., Ltd. (Shanghai, China); H2S assay kit
was purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing,
China); RhoA activity assay kit, ROCK1 activity assay kit,
and ROCK2 activity assay kit were purchased from Jiangsu
Meimian Industrial Co., Ltd. (Yancheng, China); Dulbecco’s
modified Eagle’s medium (DMEM) was purchased from HyClone (Beijing,
China).
ACh, Atr, C3TF, PPG, and Y27632 were
purchased from Sigma Chemical (St. Louis); Asp was purchased from
Solarbio (Beijing, China); KD025 was purchased from Merck (Darmstadt,
Germany); CCG-1423 was purchased from ApexBio (Houston); fura-2-acetoxymethyl
ester (Fura-2 AM) was purchased from Dojindo (Kumamoto, Japan); anti-α-SMA
antibody, anti-β-actin antibody, and anti-p-MLC antibody were
purchased from Affinity Biosciences (Changzhou, China); anti-CSE antibody
and anti-RhoA antibody were purchased from Abcam (San Francisco);
anti-3-MST antibody was purchased from Santa Cruz Biotechnology (Santa
Cruz); anti-factor VIII antibody was purchased from Shanghai Fushen
Biotechnology Co., Ltd. (Shanghai, China); H2S assay kit
was purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing,
China); RhoA activity assay kit, ROCK1 activity assay kit,
and ROCK2 activity assay kit were purchased from Jiangsu
Meimian Industrial Co., Ltd. (Yancheng, China); Dulbecco’s
modified Eagle’s medium (DMEM) was purchased from HyClone (Beijing,
China).
5
Western Blot Analysis of CPA4 Protein
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Briefly, proteins were separated by 10% SDS‐PAGE and then transferred to PVDF membranes (Millipore). The membranes were incubated with 5% skim milk and then incubated with antibody against CPA4 (1:1000, Abcam), followed by incubation with an HRP‐labelled secondary antibody (CST) and detection by chemiluminescence. An anti‐β‐actin antibody (1:1000, Affinity, USA) served as control.
6
GFRA1 Protein Expression Analysis
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Western blot analysis was performed using standard procedures. Briefly, total proteins were extracted and separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred onto a polyvinylidene difluoride (PVDF) membrane (Millipore, USA). To block nonspecific binding, the membranes were incubated with 5% skim milk powder at room temperature for one hour. The membrane was then incubated with primary antibody against GFRA1 (1:1000, CST, USA), followed by horseradish peroxidase (HRP)-labeled secondary antibody (Santa Cruz) and detected by chemiluminescence. An anti-β-actin antibody (1:1000, Affinity, USA) was used as a protein loading control.
7
Protein Expression Analysis by Western Blot
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Briefly, proteins were extracted, quantified and separated by 10% SDS-PAGE and transferred to PVDF membranes (Millipore). Then incubated with 5% skim milk at room temperature for 1 h and with primary antibody of KIF4A (1:100, Abcam, USA). Then incubated with HRP-labeled secondary antibody (CST) and detected by chemiluminescence. Anti-β-actin antibody (11,000, Affinity, USA) was used as a control.
8
Wnt Signaling Pathway Inhibition in Ovarian Cancer
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OV119 and A2780 cells were cultured with or without the addition of the Wnt–β-catenin cascade inhibitor Dkk1 (100 ng/ml, R&D Systems). Proteins of OV119 and A2780 were extracted, and then the concentration determined, fractionated by SDS-PAGE gel (10%) and transfer-embedded to PVDF membranes (Millipore, USA). Then, membranes were blocked with 5% skim milk for 1 h at room temperature. Then the membranes were inoculated with primary antibodies against E-cadherin (Cell Signaling Technology, USA; 1:1000), N-cadherin (Cell Signaling Technology; 1:1000), vimentin (Cell Signaling Technology; 1:1000), wnt-1 (Abcam, USA; 1:1000), β-catenin (Cell Signaling Technology; 1:1000), as well as c-myc (Cell Signaling Technology; 1:1000). Then, an HRP-labeled secondary antibody (Cell Signaling Technology) was added. The anti-β-actin antibody (Affinity, USA; 1:1000) was used as the control.
9
Western Blot Analysis of FKBP3
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In brief, proteins were extracted, quantified, and separated by 10% SDS-PAGE and transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, USA); then incubated with 5% skim milk at room temperature for 1 h and with primary antibody of FKBP3 (1:1,000; Abcam, USA); and then incubated with horseradish peroxidase (HRP)-labeled secondary antibody (CST) and detected by chemiluminescence. Anti-β-actin antibody (1:1,000; Affinity, USA) was used as a control.
10
Protein Expression Analysis by Western Blot
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Western blot analysis was performed using standard procedures. Briefly, total proteins were extracted and separated by 10% SDS-PAGE and transferred onto a PVDF membrane (Millipore, USA). To block nonspecific binding, the membranes were incubated with 5% skim milk powder at room temperature for one hour. The membrane was then incubated with primary antibody against FBLIM1 (1:1000, Abcam, USA), followed by HRP-labeled secondary antibody (Santa Cruz) and detected by chemiluminescence. An anti-β-actin antibody (1:1000, Affinity, USA) was used as a protein loading control.
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