Tissue samples were ground to powder in liquid nitrogen. Total proteins were extracted from tissue powder or treated cells using sodium dodecyl sulfate lysis buffer (Beyotime, Shanghai, People’s Republic of China) for 30 minutes at 4°C, and an equal amount of protein was separated using 10% polyacrylamide sodium dodecyl sulfate gels. Then, the proteins were transferred to polyvinylidene fluoride membranes (Thermo Fisher Scientific) and were probed with primary antibodies against E-cadherin (Abcam, Cambridge, UK), vimentin (GeneTex, San Antonio, TX, USA), TWIST1 (Abcam), N-cadherin (GeneTex), matrix metalloproteinase (MMP)-2 (Abcam), MMP-9 (Abcam), or GAPDH (Gene-Tex). The membranes were incubated overnight at 4°C, followed by incubation with secondary antibody peroxidase-conjugated anti-IgG (Abcam), and detected using a chemiluminescent detection system (Pierce ECL Substrate Western blot detection system; Thermo Fisher Scientific). Quantity One 4.5.0 software (Bio-Rad Laboratories Inc., Hercules, CA, USA) was used to quantify the integrated density of the protein bands.
Peroxidase conjugated anti igg
Manufactured by Abcam
Sourced in United States
Peroxidase-conjugated anti-IgG is a secondary antibody that is covalently linked to the enzyme horseradish peroxidase. This antibody is commonly used in immunoassays and immunohistochemistry techniques to detect and quantify the presence of immunoglobulin G (IgG) in samples.
Automatically generated - may contain errors
Lab products found in correlation
Sodium dodecyl sulfate lysis buffer, by Beyotime (1 mentions)
Polyvinylidene fluoride membrane, by Thermo Fisher Scientific (1 mentions)
Twist1, by Abcam (1 mentions)
Pierce ecl substrate western blot detection system, by Thermo Fisher Scientific (1 mentions)
Quantity one 4, by Bio-Rad (1 mentions)
Mmp 9, by Abcam (1 mentions)
E cadherin, by Abcam (1 mentions)
Gapdh, by GeneTex (1 mentions)
Vimentin, by GeneTex (1 mentions)
N cadherin, by GeneTex (1 mentions)
Gapdh antibody, by Merck Group (1 mentions)
Monoclonal β actin, by Merck Group (1 mentions)
Protein isolation kit, by Tiangen Biotech (1 mentions)
Cyclin e, by Santa Cruz Biotechnology (1 mentions)
Cxcr4, by Santa Cruz Biotechnology (1 mentions)
β actin, by Santa Cruz Biotechnology (1 mentions)
Cyclin d1, by Santa Cruz Biotechnology (1 mentions)
Pierce bca assay, by Thermo Fisher Scientific (1 mentions)
2 protocols using peroxidase conjugated anti igg
1
Western Blot Analysis of EMT Markers
2
Western Blot Analysis of Cell Signaling Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
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The Western blot assay used in this study was according to the following previously described methods.
Total proteins were extracted from A172 cells using a protein isolation kit (Tiangen, Beijing, China), and the protein concentrations were quanti ed through the Pierce BCA assay (Thermo Scienti c, CA, USA). Equal amounts of protein samples were separated by SDS-PAGE and then electrophoretically transferred to a polyvinylidene uoride membrane (Milipore, CA, USA). The antibodies against CDK4, Cyclin-D1, CDK2, Cyclin-E, β-actin, and CXCR4 (Santa Cruz Biotechnology, CA, USA) as the primary antibody (all obtained from Cell signaling Technology, Beverly, MA, USA) were used in the Western blot assay, and peroxidase-conjugated anti-IgG (Abcam) was also used as the secondary antibody. Monoclonal β-actin or GAPDH antibody (Sigma) was used as the control. The blots were detected using an enhanced chemiluminescent kit (Pierce, Rockford, IL, USA). The data of Western blot bands were quanti ed by Image J software.
Total proteins were extracted from A172 cells using a protein isolation kit (Tiangen, Beijing, China), and the protein concentrations were quanti ed through the Pierce BCA assay (Thermo Scienti c, CA, USA). Equal amounts of protein samples were separated by SDS-PAGE and then electrophoretically transferred to a polyvinylidene uoride membrane (Milipore, CA, USA). The antibodies against CDK4, Cyclin-D1, CDK2, Cyclin-E, β-actin, and CXCR4 (Santa Cruz Biotechnology, CA, USA) as the primary antibody (all obtained from Cell signaling Technology, Beverly, MA, USA) were used in the Western blot assay, and peroxidase-conjugated anti-IgG (Abcam) was also used as the secondary antibody. Monoclonal β-actin or GAPDH antibody (Sigma) was used as the control. The blots were detected using an enhanced chemiluminescent kit (Pierce, Rockford, IL, USA). The data of Western blot bands were quanti ed by Image J software.
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