The total protein from cultured cells and tumors were obtained with Laemmli sample buffer (Bio-Rad, Hercules, CA, USA), separated by 10% polyacrylamide gel electrophoresis, and transferred onto polyvinylidene fluoride membranes (Millipore, MA, USA). The protein membrane was incubated overnight at 4°C with primary antibodies (anti-pERK1/2, anti-total ERK1/2, anti-pMEK1/2, anti-MEK1/2, anti-pMER1/2, anti-pSTAT3, anti-STAT3, anti-pAKT, anti-AKT, and anti-GAPDH antibody, Abcam, Cambridge, MA, USA). We used the enhanced chemiluminescence (Thermo Scientific, Waltham, MA) to detect protein bands and semi-quantified the band density with an ImageJ analysis software (National Institutes of Health).
Anti total erk1 2
Manufactured by Abcam
Sourced in United States
Anti-total ERK1/2 is a primary antibody that binds to and detects the extracellular signal-regulated kinases 1 and 2 (ERK1/2) proteins. ERK1/2 are key members of the mitogen-activated protein kinase (MAPK) signaling pathway, which plays a crucial role in cellular processes such as proliferation, differentiation, and survival.
Automatically generated - may contain errors
Lab products found in correlation
Anti gapdh antibody, by Abcam (2 mentions)
Enhanced chemiluminescence, by Thermo Fisher Scientific (2 mentions)
Laemmli sample buffer, by Bio-Rad (2 mentions)
Anti p erk1 2, by Abcam (2 mentions)
Anti pmek1 2, by Abcam (2 mentions)
Anti mek1 2, by Abcam (2 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Anti akt, by Abcam (1 mentions)
Anti p akt, by Abcam (1 mentions)
Anti p stat3, by Abcam (1 mentions)
Anti stat3, by Abcam (1 mentions)
Enhanced chemi luminescence (ecl), by Merck Group (1 mentions)
Anti nrf2, by Abcam (1 mentions)
β actin, by Abcam (1 mentions)
Anti phospho erk1 2, by Cell Signaling Technology (1 mentions)
Anti phospho aktser473, by Bioworld Technology (1 mentions)
Pvdf membrane, by Roche (1 mentions)
Anti gapdh, by Abcepta (1 mentions)
Anti e cadherin, by Abcam (1 mentions)
Anti vimentin, by Abcam (1 mentions)
6 protocols using anti total erk1 2
1
Western Blot Analysis of Signaling Proteins
2
Western Blot Analysis of Lung Protein
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The protein expression in the lung tissues was evaluated through Western blot. Briefly, total protein extracts from lung tissues were separated by polyacrylamide gradient gels (12%) and transferred to polyvinylidene fluoride membranes. The membranes were incubated overnight at 4°C with primary antibodies anti‐phospho‐ERK1/2 (Cell Signalling Technology, Boston, MA), anti‐Nrf2, anti‐total ERK1/2 (Abcam Cambridge, UK), and β‐actin (1:1000 dilution). Then, the respective secondary antibodies conjugated to HRP were incubated for 1 hour, followed by 3 times of washing. The membranes were then incubated with ECL (Millipore, MA, USA) for luminescence generation. The image and grey level were analysed with Alpha Innotech and Adobe software. The protein expression level was evaluated with the β‐actin ratio.
3
Western Blot Analysis of Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The antibodies purchased were as follows: anti-GAPDH antibody, anti-phospho ERK1/2 from Cell Signaling Technology; anti-total ERK1/2 from Abcam Biotechnology. Protein lysates of cells were resolved by 10% SDS–polyacrylamide gel electrophoresis (SDS–PAGE) and transferred onto PVDF membranes. After the membranes were blocked by 5% nonfat milk, the membrane was incubated with specific antibodies as well as the secondary antibodies labeled with horseradish peroxidase and visualized by chemiluminescence [28 (link)].
4
Western Blot Analyses of Signaling Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were lysed in prechilled RIPA buffer containing protease inhibitors. The protein lysates were separated on SDS–PAGE and then transferred to PVDF membranes (Roche Diagnostics, Mannheim, Germany). The membranes were blocked for 2 h in 5% bovine serum albumin (BSA) in 1 × TBS-T (0.5% Tween-20) and incubated with the indicated primary antibodies, including anti-EHF (Abcam, Inc), anti-total-Erk1/2 (Abcam, Inc), anti-phospho-Erk1/2 (Epitomics, Inc), anti-phospho-AktSer473 (Bioworld Technology, co, Ltd), anti-total-Akt (Bioworld Technology, co, Ltd), anti-HER2 (Sino Biological, Inc), anti-HER3 (Sino Biological, Inc), anti-HER4 (Sino Biological, Inc), anti-E-cadherin (Epitomics, Inc), anti-Vimentin (Epitomics, Inc) and anti-GAPDH (Abgent, Inc). The membranes were then incubated with species-specific HRP-conjugated secondary antibodies from ZSGB-BIO, and immunoblotting signals were visualized using the Western Bright ECL detection system (Advansta, CA).
5
Immunoblot Analysis of Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The total protein from cultured cells and tumors were obtained with Laemmli sample buffer (Bio-Rad, Hercules, CA, USA) and separated by 10% polyacrylamide gel electrophoresis, followed by transferring onto polyvinylidene uoride membranes (Millipore, MA, USA). The protein membrane was incubated with primary antibodies (Anti-pERK1/2, anti-total ERK1/2, Anti-pMEK1/2, Anti-MEK1/2, and Anti-GAPDH antibody, Abcam, Cambridge, MA, USA) overnight at 4 °C. We used the enhanced chemiluminescence (Thermo Scienti c, Waltham, MA) to detect protein bands and semi-quanti ed the band density with an ImageJ analysis software (National Institutes of Health).
6
Western Blot Analysis of Signaling Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A total of 10 6 cells for each treatment were pelleted and lysed in RIPA lysis buffer (Sigma) supplemented with protease inhibitor cocktail (Roche). Cell lysates were centrifuged at 12000g, 4 °C for 15 min and protein concentration was measured using bicinchoninic acid assay kit (ThermoFisher). Proteins were separated by SDS-PAGE in reducing conditions and transferred to nitrocellulose membrane (GE Healthcare Life Sciences). After saturation with 5% milk, the membrane was incubated overnight at 4 °C with the appropriate primary antibody: anti-Phospho ERK1/2 1:5000, anti-total ERK 1/2 (1:10000), anti-Phospho Akt (1:5000), anti-total Akt (1:10000), anti-Phospho FAK (1:1000), and anti-total FAK (1:1000) (Abcam), anti-APOL1 (1/1000), anti-APOL2 (1/ 2000), anti-APOL3 (1/2500) and anti-APOL6 (1/500) (sigma), washed, then incubated with (HRP) conjugated secondary antibody for 1 h at room temperature and revealed using the ECL substrate (PerkinElmer). Monoclonal anti-β-actin antibody (1/80000; Sigma) was used for protein loading control. Densitometric analysis was performed using ImageJ.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!